Cyclodextrin dimers and their use

Abstract

This invention provides a compound for preferentially solubilizing and removing 7-ketocholesterol (7KC), which may be a causative agent of various diseases. [Solution] Cyclodextrin (CD) dimers, CD compositions, and their uses are disclosed. CD dimers and their compositions may be useful for a variety of purposes, including targeting 7KC. The design and testing of CD dimers are described herein. Exemplary CD dimers include heterodimers, homodimers, or asymmetric dimers.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the interests of U.S. Patent No. 63 / 048,824, U.S. Patent No. 63 / 048,886, and U.S. Patent No. 63 / 048,941, filed on July 7, 2020, and is a continuation of U.S. Patent No. 16 / 733,945, filed on January 3, 2020, which in turn claims the interests of U.S. Patent No. 62 / 787,869, filed on January 3, 2019, and U.S. Patent No. 62 / 850,334, filed on May 20, 2019, each of which is incorporated herein by reference in its entirety. [Background technology]

[0002] 7-Ketocholesterol (7KC) is an oxysterol produced by a non-enzymatic reaction between oxygen radicals and cholesterol. 7KC may be formed in the body or ingested through food, but it is considered potentially toxic and of no use in humans and other eukaryotes. Like cholesterol, 7KC is found in atherosclerotic plaques. It is the most abundant non-enzymatically produced oxysterol in atherosclerotic plaques and may be a contributing factor to atherosclerosis and age-related diseases, lysosomal storage disorders such as Niemann-Pick disease type C (NPC), heart disease, cystic fibrosis, liver damage and failure, and complications of hypercholesterolemia. In individuals with hypercholesterolemia, 7KC can diffuse across cell membranes, in which case it affects receptor and enzyme function. The high rate of dementia in hypercholesterolemia has been associated with 7KC accumulation. In the liver, 7KC affects tissue fenestration and porosity, which increase with age. 7KC also promotes the migration of cytoplasmic NADPH oxidase components to neutrophil (leukocyte) membranes and enhances rapid reactive oxygen species production. Elevated 7KC levels have also been linked to the pathogenesis of other age-related diseases, such as age-related macular degeneration (AMD - atrophic type), Alzheimer's disease, and lysosomal storage disorders like Niemann-Pick disease type C (NPC). Oxysterols, including 7KC, are also involved in elevated free radical levels, which in turn affect lipid circulation in cystic fibrosis. Increased free radicals induced by oxysterols like 7KC are thought to be involved in apoptosis, cytotoxicity, endothelial dysfunction, and the regulation of enzymes involved in inflammation and fatty acid metabolism.

[0003] 7KC is formed from a non-enzymatic reaction between oxygen radicals and cholesterol, which suggests that 7KC formation may not be beneficial. In fact, 7KC is thought to enhance free radical production throughout the body, but this is of particular concern in cardiovascular tissue. Free radicals affect cellular and enzymatic reactions that are important for cholesterol-mediated tissue damage, and this is especially critical in these tissues. Free radicals are thought to enhance inflammation in vascular structures. 7KC is thought to cause mitochondrial and lysosome dysfunction by disrupting the function of cell and organelle membranes, and is thought to be involved in increasing the frequency of foam cell formation from macrophages in atherosclerotic plaques. While the scavenging function of these macrophages is expected to help alleviate plaque, if they are filled with cholesterol and oxysterols, they may not, and instead become part of the plaque.

[0004] Cyclodextrins (CDs) are cyclic oligosaccharides composed of a 6(αCD), 7(βCD), or 8(γCD) sugar ring (Figure 1A). Alpha-CD, beta-CD, and gamma-CD are the most common forms and have many applications related to medicine, industry, consumer goods, and food. CDs have been used in a variety of applications, such as dietary fiber as a food additive. CDs have also been used in pharmaceutical compositions as aerosolizing agents and, typically in combination with active pharmaceutical ingredients, as excipients for small hydrophobic drugs.

[0005] Previous studies have shown that two βCDs can be covalently bonded head-to-head, significantly improving their ability to form inclusion complexes with target molecules such as 7KC. Furthermore, it has been shown that αCD, when complexed with 1,12-diaminododecane, dimerizes non-covalently in a head-to-head manner. The structure of this guest molecule includes a long aliphatic chain. [Overview of the project]

[0006] This disclosure describes additional CD dimers that may be useful for a variety of purposes, including the targeting of 7KC. The design and testing of CD dimers are described herein. Examples of CD dimers include heterodimers (preferably comprising αCD and βCD), homodimers (preferably comprising two αCD or two βCD having the same substituents on each monomer), or asymmetric dimers (e.g., having two CD monomers having different substituent combinations).

[0007] The inventors hypothesized that, with respect to CD heterodimers, the smaller ring structure of αCD (compared to βCD and γCD) favorably interacts with the 7KC tail group. In particular, the inventors propose that a linked CD dimer composed of one αCD and one βCD selectively and asymmetrically targets both portions of the guest molecule (βCD forms a complex with the head group, while αCD encapsulates the tail). Other guest molecules having similar structures can be encapsulated in the same manner. None of these CD monomers may have substituents added to alter solubility, either one may have such substituents, or both may have such substituents.

[0008] Furthermore, we propose unusual and / or novel substituents that can enhance the target specificity of these molecules in addition to the CD dimers. Examples of CD dimers disclosed herein include substituents that can increase the affinity and / or specificity of these CD dimers to target molecules, such as 7KC, cholesterol, and other sterols. Certain substituted CDs described herein are predicted to interact strongly with the carbonyl group of 7KC. Since cholesterol does not have a carbonyl group, we hypothesize that such substitutions will produce a more significant specificity to 7KC than to cholesterol.

[0009] In one embodiment, the present disclosure provides alpha-beta heterodimers of CD, which are variously substituted, such as a combination of one αCD and one βCD monomer, and which can exhibit enhanced binding properties.

[0010] CD heterodimers with an αCD bonded to a βCD have a smaller αCD cavity, which more effectively encapsulates the 7KC tail group and reduces the likelihood of binding with other bulky hydrophobic molecules. Both αCD and βCD can be substituted with various chemical groups to adjust the subunit's affinity for the target head or tail group of the target molecule.

[0011] In another embodiment, the disclosure provides CD heterodimers in which alkyl groups are used as substituents. Because alkyl groups are more hydrophobic than previously demonstrated charged and polar substituents, they expand the hydrophobic cavities of one or both subunits. This creates a better environment for encapsulating the tail groups of 7KC, cholesterol, and other sterols having long aliphatic chains.

[0012] Furthermore, this disclosure describes the design and testing of various asymmetric dimers of CD, including (2-hydroxypropyl)-βCD (HPβCD) dimers, methyl-βCD (MeβCD) dimers, succinyl-βCD (SUCCβCD) dimers, sulfobutyl-βCD (SBβCD) dimers, quaternary ammonium-βCD (QAβCD) dimers, and the like. Asymmetric dimers include combinations of two different CD monomers. The exemplary asymmetric dimers of this disclosure exhibit enhanced binding properties.

[0013] The exemplary asymmetric dimers of this disclosure may be useful for targeting 7KC. For example, the asymmetric dimer may include, for example, two specifically substituted monomers, such as unmodified βCD linked to HPβCD, or SBβCD linked to MeβCD. Although not intended to be theoretically limiting, the affinity and specificity of the CD asymmetric dimer to the guest may change depending on the type of substitution, and asymmetric substitution of the dimer may make it more specific to the target head or tail group. In exemplary embodiments, substitution at one position, for example, the C6 position, is expected to produce a more uniform product at synthesis and (although not intended to be theoretically limiting) extend the CD cavity and enhance the ability to solubilize hydrophobic molecules such as sterols (e.g., 7KC).

[0014] Furthermore, we describe substituents that can be added to asymmetric CD dimers that may enhance the target specificity of these molecules. Some of the substitutions and substitution patterns described herein are expected to promote specific or preferential binding of 7KC. Although not intended to be limited by theory, it is thought that these substitutions may produce more pronounced specificity for 7KC than cholesterol, since cholesterol does not have a carbonyl group.

[0015] Furthermore, we discuss the substitution of opposite charges between the two CD subunits of the asymmetric dimer, namely a positive charge substitution of one monomer and a negative charge substitution of the other monomer. Although not intended to be limited by theory, such substitutions are predicted to promote the stability of the CD-labeled complex by providing an electrostatic attraction between the two CD subunits of the asymmetric dimer, and / or provide specificity to target molecules containing highly polar regions that can interact with the charged moiety.

[0016] Embodiments of the present invention provide compositions and methods for treating or preventing atherosclerosis and other age-related diseases. 7KC is the most abundant non-enzymatically produced oxysterol in atherosclerotic plaques and is thought to be a contributing factor to the development of atherosclerosis. Treatment using the asymmetric CD dimer of the present invention is expected to be beneficial in preventing and / or improving atherosclerotic plaque formation.

[0017] In another embodiment, the present disclosure provides a method for producing asymmetric CD dimers with specificity to further small molecules. An exemplary method is carried out by first producing a CD dimer nucleus having a specific (possibly asymmetric) structure to which specificity is conferred during synthesis. Then, arbitrary substitutions can be made while maintaining the high affinity transmitted by the CD dimer nucleus to create the specificity to the hydrophobic molecules described above. This specificity can be further modified with different linkers.

[0018] Furthermore, this disclosure describes the design and testing of various homodimers (CDs) of CD, including HPβCD dimers, methyl-βCD dimers, succinyl-βCD dimers, sulfobutyl-βCD dimers, quaternary ammonium-βCD dimers, and others. This disclosure describes dimers consisting of combinations of two CD monomers. Exemplary homodimers exhibit enhanced binding properties to target molecules, including 7KC, cholesterol, and other sterols, including exemplary homodimers with increased specificity for 7KC than for cholesterol.

[0019] Exemplary embodiments provide the use of the CD dimers of the present disclosure (e.g., heterodimers, homodimers, or asymmetric dimers) in compositions, and methods for treating diseases associated with and / or exacerbated by the accumulation of 7KC, including atherosclerosis, AMD, arteriosclerosis, coronary artery atherosclerosis due to coronary artery calcification lesions, heart failure (all stages), Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, Huntington's disease, vascular dementia, multiple sclerosis, Smith-Lemle-Oppitz syndrome, Examples of conditions that may be affected include infantile neuronal ceroid lipofuscinosis, lysosomal acid lipase deficiency, cerebral tendon xanthomatous cerebrospinal fluid, X-linked adrenoleukodystrophy, sickle cell anemia, Niemann-Pick disease type A, Niemann-Pick disease type B, Niemann-Pick disease type C, Gaucher disease, Stargardt disease, idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, cystic fibrosis, liver injury, liver failure, non-alcoholic steatohepatitis, non-alcoholic fatty liver disease, irritable bowel syndrome, Crohn's disease, ulcerative colitis, and / or hypercholesterolemia or dementia associated with hypercholesterolemia. Preferred CD dimers, i.e., heterodimers, homodimers, or asymmetric dimers, are selective for 7KC (compared to cholesterol). Preferably, the above CD dimers preferentially solubilize 7KC while minimizing or avoiding potentially harmful or toxic effects that may result from excessive removal of cholesterol.

[0020] Exemplary embodiments of the present invention provide the use of CD (e.g., HPα-βCD, Meα-βCD, SUCCα-βCD, QAα-βCD, or SBα-βCD) dimers for solubilization and / or removal of 7KC which can be performed in vitro or in vivo.

[0021] In exemplary embodiments, the above-mentioned CD (e.g., HPα-βCD, MEα-βCD, SUCCα-βCD, QAα-βCD, or SBα-βCD) dimers exhibit higher binding affinity and / or solubilization of 7KC than cholesterol. While the specificity of 7KC to cholesterol is most pronounced at near-saturated concentrations, solubilization of both sterols at higher concentrations can approach 100%. This specificity allows for the use of such CD dimers to preferentially solubilize and remove 7KC.

[0022] The term "pharmaceutically acceptable" is used herein to refer to a compound, material, composition, and / or dosage form that, within the bounds of sound medical judgment, is suitable for entry into a living organism or living biological tissue, and preferably, is free from significant toxicity, irritation, or allergic reactions. The present invention comprises a method comprising administering a CD dimer to a patient, wherein the CD dimer is contained within a pharmaceutical composition. The pharmaceutical compositions of the present invention are formulated with pharmaceutically acceptable carriers, excipients, and other agents that provide suitable transport, delivery, tolerance, etc. Numerous suitable formulations can be found in formularies known to pharmaceutical chemists, e.g., Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa. Examples of these formulations include powders, pastes, ointments, gels, waxes, oils, lipids, lipid (cationic or anionic)-containing vesicles (e.g., LIPOFECTIN®), DNA complexes, anhydrous absorbent pastes, oil-in-water and water-in-oil emulsions, carbowax emulsions (polyethylene glycol of various molecular weights), semi-solid gels, and carbowax-containing semi-solid mixtures. (See also Powell [et al.], J. Pharm. Sci. Technol., 52:238-311, (1998)).

[0023] When used herein, the term “pharmaceutically acceptable carrier” generally refers to a pharmaceutically acceptable composition useful for introducing an active agent into the body, such as a liquid or solid filler, diluent, excipient, manufacturing aid (e.g., lubricant, magnesium talc, calcium stearate or zinc stearate, or stearic acid), or solvent encapsulation material. Each carrier must be “acceptable” in the sense that it is compatible with the other components of the formulation and is not harmful to the patient. Examples of suitable aqueous and non-aqueous carriers usable in the pharmaceutical compositions of the present invention include, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.), vegetable oils (e.g., olive oil), and organic esters for injection (e.g., ethyl oleate), as well as suitable mixtures thereof. Adequate fluidity can be maintained, for example, by the use of a coating material such as lecithin, by maintaining the required particle size in the case of dispersion, and by the use of a surfactant.

[0024] Other examples of materials that can function as pharmaceutically acceptable carriers include: (1) sugars, e.g., lactose, glucose, and sucrose; (2) starches, e.g., corn starch and potato starch; (3) cellulose and its derivatives, e.g., sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, e.g., cocoa butter and suppository wax; (9) oils, e.g., peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; (10) glycols. Examples include (11) propylene glycol, polyols such as glycerin, sorbitol, mannitol, and polyethylene glycol, (12) esters such as ethyl oleate and ethyl laurate, (13) agar, (14) buffers such as magnesium hydroxide and aluminum hydroxide, (15) alginic acid, (16) pyrogen-free water, (17) isotonic saline, (18) Ringer's solution, (19) ethyl alcohol, (20) pH buffer, (21) polyesters, polycarbonates, and / or polyanhydrides, and (22) other non-toxic and suitable substances used in pharmaceutical formulations.

[0025] Various auxiliary agents such as wetting agents, emulsifiers, lubricants (e.g., sodium lauryl sulfate and magnesium stearate), colorants, release agents, coating agents, sweeteners, flavorings, preservatives, and antioxidants may also be present in the pharmaceutical composition. Some examples of pharmaceutically acceptable antioxidants include (1) water-soluble antioxidants such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfite, sodium metabisulfite, and sodium sulfite; (2) oil-soluble antioxidants such as ascorbic acid palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, and alpha-tocopherol; and (3) metal chelating agents such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, and phosphoric acid. In some embodiments, the pharmaceutical formulation includes excipients selected from, for example, cellulose, liposomes, micellar-forming agents (e.g., bile acids), and polymer carriers, such as polyesters and polyanhydrides. The suspension may contain, in addition to the active compound, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxylate, bentonite, agar, and tragacanth, as well as mixtures thereof. Various antibacterial and antifungal agents, such as parabens, chlorobutanol, and phenolsorbic acid, can be included to reliably prevent microbial action of the active compound. It may also be desirable to include isotonic agents such as sugars and sodium chloride in the composition. Furthermore, the inclusion of absorption-delaying agents such as aluminum monostearate and gelatin may provide sustained absorption of the injectable pharmaceutical form.

[0026] The pharmaceutical formulations of the present invention can be prepared by any method known in the pharmaceutical art. The amount of the active ingredient (i.e., a CD dimer such as HPβCD dimer or another CD dimer of this disclosure) that can be combined with a carrier material to make a single dosage form varies depending on the host being treated and the specific mode of administration. The amount of the active ingredient that can be combined with a carrier material to make a single dosage form is generally the amount of compound that produces the therapeutic effect. The amount of the active compound may range from about 0.1 to 99.9 percent, more typically from about 80 to 99.9 percent, and more typically from about 99 percent. The amount of the active compound may range from about 0.1 to 99 percent, more typically from about 5 to 70 percent, and more typically from about 10 to 30 percent. In exemplary embodiments, dosage forms for intravenous administration are provided, which are aqueous solutions with a concentration of 0.5% to 0.001%, for example, 0.12% to 0.0105%, for example, about 0.01% (W / V). In exemplary embodiments, dosage forms for intravenous administration are provided, which are aqueous solutions with concentrations of 2.5% to 0.25%, for example, 2% to 0.5%, for example, about 1% (W / V). In exemplary embodiments, the dosage form provides intravenous administration of up to 500 mL of a 1% (W / V) solution, which results in a maximum dose of 5 grams.In further exemplary mechanisms, the total concentration can be up to approximately 60%(w / v) or approximately 50%(w / v), for example, approximately 5%(w / v), approximately 10%(w / v), approximately 15%(w / v), approximately 20%(w / v), approximately 25%(w / v), approximately 30%(w / v), approximately 35%(w / v), approximately 40%(w / v), approximately 45%(w / v), approximately 50%(w / v), approximately 55%(w / v). ), or approximately 60% (w / v); or at least approximately 5% (w / v), at least approximately 10% (w / v), at least approximately 15% (w / v), at least approximately 20% (w / v), at least approximately 25% (w / v), at least approximately 30% (w / v), at least approximately 35% (w / v), at least approximately 40% (w / v), at least approximately 45% (w / v), and at least Also approximately 50% (w / v), or at least approximately 55% (w / v), or approximately 1% (w / v) to 60% (w / v), 5% (w / v) to 55% (w / v), 10% (w / v) to 50% (w / v), 15% (w / v) to 45% (w / v), 20% (w / v) to 40% (w / v), 25% (w / v) to 35% (w / v), or approximately 30% (w / v); or Dosage forms are provided that contain one or more CDs in a maximum of approximately 10% (w / v), 15% (w / v), 20% (w / v), 25% (w / v), 30% (w / v), 35% (w / v), 40% (w / v), 45% (w / v), 50% (w / v), 55% (w / v), or 60% (w / v). The CD dosage form can be formulated for administration to patients, for example, parenteral administration, preferably intravenous administration, and such administration optionally includes dilution to pre-administration concentrations of approximately 5% (w / v), approximately 10% (w / v), approximately 15% (w / v), approximately 20% (w / v), approximately 25% (w / v), approximately 30% (w / v), or approximately 35% (w / v).

[0027] In exemplary embodiments, a patient may be administered 1 mg to 10 g of CD dimers, for example, 10 mg to 1 g or 100 mg to 500 mg. In exemplary embodiments, a patient may be administered about 400 mg of CD dimers. In exemplary embodiments, a patient may be administered 1 to 10 g of CD dimers, for example, about 2 g, about 3 g, about 4 g, or about 5 g. In exemplary embodiments, a patient may be administered 50 mg to 5 g of CD dimers, for example, 100 mg to 2.5 g, 100 mg to 2 g, 250 mg to 2.5 g, for example, about 1 g.

[0028] In exemplary embodiments, a single dosage form is provided which may contain the aforementioned amounts of CD dimers, may be packaged for individual administration, and may optionally further contain a pharmaceutically acceptable carrier or excipient. The total amount of CD dimers in the single dosage form may be as presented above, for example, 1 mg to 10 g, e.g., 10 mg to 1 g, 100 mg to 500 mg, 1 to 10 g of CD dimers, 50 mg to 5 g, 100 mg to 2.5 g, 100 mg to 2 g, 250 mg to 2.5 g, e.g., about 1 g, 2 g, about 3 g, about 4 g, or about 5 g.

[0029] Formulations of the present invention suitable for oral administration may be in the form of capsules, cachets, pills, tablets, sweetened tablets (using flavoring agents, usually sucrose and acacia or tragacanth), powders, granules, or solutions or suspensions in aqueous or non-aqueous liquids, or oil-in-water or water-in-oil liquid emulsions, or elixirs or syrups, or pastils (using inert agents such as gelatin and glycerin, or sucrose and acacia), and / or mouthwashes, each containing a predetermined amount of the compound of the present invention as the active ingredient. The active compound may also be administered as a bolus, lick, or paste.

[0030] The preparation methods for these formulations or compositions generally involve the step of mixing the compound of the present invention with a carrier and optionally one or more auxiliary agents. In the case of solid dosage forms (e.g., capsules, tablets, pills, powders, granules, lozenges, etc.), the active compound can be mixed with a pulverized solid carrier and molded, typically by pelletizing, tableting, granulating, powdering, or coating. Generally, examples of solid carriers include sodium citrate or dicalcium phosphate and / or any of the following: (1) Fillers or bulking agents, e.g., starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; (2) Binders, e.g., carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and / or acacia; (3) Humectants, e.g., glycerol; (4) Disintegrants, e.g., agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; (5) Dissolution retarders, e.g., paraffin; (6) Absorption enhancers, e.g., quaternary ammonium compounds and ammonium compounds. (7) surfactants, such as poloxamer and sodium lauryl sulfate; (8) wetting agents, such as cetyl alcohol, glycerol monostearate, and nonionic surfactants; (9) absorbents, such as kaolin and bentonite clay; (10) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, zinc stearate, sodium stearate, stearic acid, and mixtures thereof; (11) colorants; and (22) release control agents, such as crospovidone or ethylcellulose. In the case of capsules, tablets, and pills, the pharmaceutical composition may also contain buffers. Similar types of solid compositions may also be used as fillers in soft and hard shell gelatin capsules, using excipients such as lactose and high molecular weight polyethylene glycol.

[0031] Tablets may be manufactured by compression or molding with one or more auxiliary components as optional. Compressed tablets may be prepared using binders (e.g., gelatin or hydroxypropyl methylcellulose), lubricants, inert diluents, preservatives, disintegrants (e.g., sodium starch glycolate or cross-linked sodium carboxymethylcellulose), surfactants, or dispersants.

[0032] Tablets and other solid dosage forms of activators, such as capsules, pills, and granules, can optionally be surface-treated or prepared using coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical manufacturing field. These dosage forms can also be formulated to release the active ingredient in a sustained or controlled manner, for example, by using hydroxypropyl methylcellulose in varying proportions to achieve a desired release profile, along with other polymer matrices, liposomes, and / or microspheres. Alternatively, the dosage forms may be formulated for rapid release, for example, by lyophilization.

[0033] In general, dosage forms need to be sterilized. For this purpose, dosage forms can be sterilized, for example, by filtration with a bacterial collection filter, or by incorporating a sterilizing agent in the form of a sterile solid composition that can be dissolved in sterile water or some other sterile injection medium immediately before use. Pharmaceutical compositions may contain opacifiers and may release the active ingredient(s) in a manner that is selectively delayed only in or preferentially in specific parts of the gastrointestinal tract. Examples of embedding compositions that can be used include polymers and waxes. The active ingredient(s) may also be in a microencapsulated form together with one or more of the excipients described above, where appropriate.

[0034] Liquid dosage forms are typically pharmaceutically acceptable emulsions, microemulsifies, solutions, suspensions, syrups, or elixirs of the active ingredient. In addition to the active ingredient, liquid dosage forms may include, for example, water or other solvents, solubilizers and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerol, tetrahydrofuryl alcohol, polyethylene glycol and sorbitan fatty acid esters, and mixtures thereof, which are commonly used in the art.

[0035] Dosage forms specifically intended for topical or transdermal administration may be, for example, in the form of powders, sprays, ointments, pastes, creams, lotions, gels, solutions, or patches. Ophthalmic formulations, such as ophthalmic ointments, powders, solutions, etc., are also intended herein. The active compound may be mixed under sterile conditions with a pharmaceutically acceptable carrier and any preservatives, buffers, or propellants as needed. In addition to the active compound of the present invention, topical or transdermal dosage forms may contain one or more excipients, such as animal and vegetable fats, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicones, bentonite, silicic acid, talc, and zinc oxide, and mixtures thereof. Sprays may also contain common propellants, such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, such as butane and propane.

[0036] With regard to the object of the present invention, transdermal patches may offer the advantage of being able to deliver the compounds of the present invention to the body in a controlled manner. Such dosage forms can be prepared by dissolving or dispersing the compounds in a suitable culture medium. Absorption enhancers may also be included to increase the flow of the compounds across the skin. Such flow rates can be controlled by either providing a rate-controlled membrane or dispersing the compounds in a polymer matrix or gel.

[0037] A pharmaceutical composition of the present invention suitable for parenteral administration generally comprises one or more compounds of the present invention in combination with one or more pharmaceutically acceptable sterile isotonic aqueous or nonaqueous solutions, dispersions, suspensions, or emulsions, or a sterile powder that can be reconstituted into a sterile injection solution or dispersion before use, the sterile powder of which may contain sugars, alcohols, antioxidants, buffers, bacteriostatic agents, or solutes that make the formulation isotonic with the blood of the target recipient.

[0038] In some cases, it may be desirable to delay drug absorption from subcutaneous or intramuscular injection in order to prolong the drug's effect. This can be achieved by using liquid suspensions of crystalline or amorphous materials with poor water solubility. The rate of drug absorption may depend on the rate of dissolution and, therefore, on the crystal size and crystal form. Alternatively, the delay of absorption of parenterally administered dosage forms can be achieved by dissolving or suspending the drug in an oily vehicle.

[0039] Depot injection formulations can be prepared by forming a microencapsulation matrix in which the active compound is contained within a biodegradable polymer such as polylactide-polyglycolide. The drug release rate can be controlled depending on the ratio of the drug to the polymer and the properties of the specific polymer used. Other examples of biodegradable polymers include poly(orthoester) and poly(anhydride). Depot injection formulations can also be prepared by capturing the drug in liposomes or microemulsifies that are compatible with body tissues.

[0040] Pharmaceutical compositions may be in the form of microemulsions. In the form of microemulsions, the bioavailability of the active ingredient may be improved. See (Dorunoo [et al.], Drug Development and Industrial Pharmacy, 17(12):1685-1713 (1991)) and (Sheen [et al.], J. Pharm. Sci., 80(7):712-714, (1991)). The contents of these documents are incorporated herein by reference in their entirety.

[0041] The pharmaceutical composition may contain micelles formed from the compound of the present invention and at least one amphiphilic carrier, in which case the micelles have an average diameter of less than about 100 nm. In some embodiments, the micelles have an average diameter of less than about 50 nm, or an average diameter of less than about 30 nm, or an average diameter of less than about 20 nm.

[0042] Although any suitable amphiphilic carrier is considered herein, amphiphilic carriers are generally those that have been approved as inactive pharmaceutical components and are capable of both dissolving the compounds of the present invention and microemulsifying them when the solution comes into contact with a complex aqueous phase (e.g., those found in living biological tissues) in a subsequent step. Typically, amphiphilic components that meet these requirements have an HLB (hydrophilic-to-lipophilic balance) value of 2 to 20 and their structure contains linear aliphatic radicals in the C-6 to C-20 range. Some examples of amphiphilic agents include polyethylene glycolated aliphatic glycerides and polyethylene glycols.

[0043] Particularly preferred amphiphilic carriers are saturated and monounsaturated polyethylene glycolated fatty acid glycers, for example, those obtained by fully or partially hydrogenating various vegetable oils. These oils may advantageously consist of tri, di, and mono fatty acid glycers, as well as di and monopolyethylene glycol esters of the corresponding fatty acids. Particularly preferred fatty acid compositions include 4%-10% capric acid, 3%-9% capric acid, 40%-50% lauric acid, 14%-24% myristic acid, 4%-14% palmitic acid, and 5%-15% stearic acid. Another useful class of amphiphilic carriers includes partially esterified sorbitan and / or sorbitol using saturated or monounsaturated fatty acids (SPAN series) or their corresponding ethoxylated analogs (TWEEN series). Commercially available amphiphilic carriers are particularly intended, and examples of such carriers include the Gelucire® series, Labrafil®, Labrasol®, or Lauroglycol®, PEG-monooleate, PEG-dioleate, PEG-monolaurate and dilaurate, lecithin, and polysorbate 80.

[0044] CD (e.g., HPβCD as disclosed herein or another CD) dimers may be administered by any suitable means. Preferred routes of administration include parenteral (e.g., subcutaneous, intramuscular, or intravenous), topical, transdermal, oral, sublingual, or buccal. Such administration may also be performed ocularly (e.g., in the form of eye drops), intravitreously, posterior orbitally, subretinally, or subsclerally, which may be preferred in cases of ocular diseases such as AMD.

[0045] The CD (e.g., HPβCD as disclosed herein or another CD) dimer may be administered to a subject or used in vitro, for example, by applying it to cells or tissues isolated from an animal. The cells or tissues can then be introduced into a subject, whether from the subject from which the cells or tissues were isolated or from another individual, the other individual preferably of the same species.

[0046] The subject receiving treatment (i.e., the patient) is typically an animal, generally a mammal, preferably a human. The subject may also be a non-human animal, such as all vertebrates, e.g., mammals, as well as non-human primates, sheep, dogs, cats, cattle, horses, chickens, amphibians, and reptiles. In some embodiments, the subject is livestock, e.g., cattle, pigs, sheep, poultry, and horses, or companion animals, e.g., dogs and cats. The subject may be genetically male or female. The subjects may be of any age, for example, the elderly (generally at least 60, 70, or 80 years old, or older), subjects in the transitional age from adulthood to old age, adults, subjects in the transitional age from pre-adulthood to adulthood, and pre-adults including young people (e.g., from 13 to 16, 17, 18, or 19 years old), children (generally under 13 years old or pre-adolescent), and infants. Furthermore, the subjects can be of any racial group or genotype. Some examples of human racial groups include the Caucasian, Asian, Latin American, African, African American, Native American, Semitic, and Pacific Islander peoples. The method of the present invention may be more suitable for certain racial groups, for example, the Caucasian, particularly Northern European, and Asian racial groups.

[0047] This disclosure includes further substitution of the dimerized CD described herein (e.g., HPβCD or another CD). Chemical modification may be performed before or after dimerization. Chemical modification of CD can be performed directly on the unmodified beta-CD ring by reacting a properly functionalized CD with a chemical reagent (nucleophile or electrophile) (Adair-Kirk [et al.], Nat. Med., 14(10):1024-5, (2008)); (Khan, [et al.], Chem. Rev., 98(5):1977-1996, (1998)). To date, more than 1,500 CD derivatives have been prepared by chemical modification of unmodified CD. CD can also be prepared by de novo synthesis starting from an oligopyranoside linked to glucopyranose. Such synthesis can be achieved using various chemical reagents or biological enzymes, such as CD transglycosylase. An overview of chemically modified CDs as drug carriers in drug delivery systems is provided, for example, in (Stella, [et al.], Toxicol. Pathol., 36(1):30-42, (2008)), the disclosure of which is incorporated herein by reference in its entirety. U.S. Patents 3,453,259 and 3,459,731 describe electrically neutral CDs, the disclosures of which are incorporated herein by reference in their entirety. Other derivatives include cationic CDs disclosed in U.S. Patent 3,453,257; insoluble crosslinked CDs disclosed in U.S. Patent 3,420,788; and anionic CDs disclosed in U.S. Patent 3,426,011, all of which are incorporated herein by reference in their entirety. Among the anionic CD derivatives, carboxylic acids, phosphorous acids, phosphinoic acids, phosphonic acids, phosphoric acids, thiophosphonic acids, thiosulfinic acids, and sulfonic acids have been added to the parent CD, as disclosed, for example, in U.S. Patent No. 3,426,011. Sulfoalkyl ether CD derivatives are also described, for example, in U.S. Patent No. 5,134,127, which is incorporated herein by reference in its entirety.In some embodiments, cyclic oligosaccharides can have two or more monosaccharide units replaced by a triazole ring, which can be synthesized by azido-alkyne hysgen cycloaddition ((Bodine, [et al.], J.Am.Chem.Soc., 126(6):1638-9, (2004)).

[0048] The dimerized CDs of this disclosure are linked by linkers. Methods available for linking the CD subunits and linkers are described in the examples. Further methods for linking the CD subunits and linkers are known in the art (Georgeta [et al.], J. Bioact. Compat. Pol., 16:39-48. (2001)), (Liu [et al.], Acc. Chem. Res., 39:681-691. (2006)), (Ozmen [et al.], J. Mol. Catal. B-Enzym., 57:109-114. (2009)), (Trotta [et al.], Compos. Interface, 16:39-48. (2009)), each of which is incorporated herein by reference in whole. For example, a linker group containing a moiety that reacts with a hydroxyl group (e.g., a carboxyl group that can be activated by carbodiimide) can be reacted with CD to form a covalent bond. In another example, one or more hydroxyl groups of CD can be activated by a known method (e.g., tosylation) and reacted with a reactive group of the linker (e.g., an amino group).

[0049] Generally, a linker initially has two reaction groups that react with and bind to each CD monomer. In one embodiment, the linker is first bound to the CD to produce a linker-CD compound, which is then isolated. The remaining reaction group of the linker in the linker-CD compound is then reacted with a second CD. The second reaction group of the linker may be protected when the first reaction group reacts, but this protection is not necessary if the first reaction group of the linker reacts with the two molecules differently. The linker can react with both molecules simultaneously and link them together. In another embodiment, the linker may have additional reaction groups for linking with other molecules.

[0050] Several linkers are known in the art. Such linkers can be used to link any of the various groups together, when the various groups possess or have been introduced with functional groups that can react with the reactive linker to link. Some groups that can react with a double reactive linker include amino, thiol, hydroxyl, carboxyl, ester, and alkyl halide groups. For example, cyclic oligosaccharides and polysaccharides can be linked using amino-amino coupling reagents when each group to be linked possesses at least one amino group. Some examples of amino-amino coupling reagents include diisocyanate, alkyl dihalides, dialdehydes, disuccinimidyl suberate (DSS), disuccinimidyl tartrate (DST), and disulfosuccinimidyl tartrate (sulfo-DST), all of which are commercially available. In another embodiment, an amino-thiol coupling agent can be used to link a thiol group of one molecule to an amino group of another molecule. Some examples of amino-thiol coupling reagents include succinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-carboxylic acid (SMCC) and sulfosuccinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-carboxylic acid (sulfo-SMCC). In yet another embodiment, a thiol-thiol coupling agent can be used to link groups having at least one thiol group.

[0051] In some embodiments, the linker is so small that its length is only one atom (e.g., --O--, --CH2--, or --NH-- linkage), or two or three atoms (e.g., amide, ureid, carbamate, ester, carbonate, sulfone, ethylene, or trimethylene linkage). In other embodiments, the linker provides a higher degree of freedom of motion by having an atomic length of at least 4, 5, 6, 7, or 8 atoms, up to a maximum of, for example, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, or 30 atoms. Preferred linker lengths are 2 to 12 atoms, or 4 to 8 atoms. In exemplary embodiments, the linker is a C4 alkyl, which may be unsubstituted. In exemplary embodiments, the linker contains a triazole.

[0052] Atherosclerosis

[0053] The exemplary CD dimers described herein are useful for the prevention or treatment of diseases such as atherosclerosis. A combination of a CD dimer and one or more activators, such as those described herein (e.g., antihyperlipidemic agents such as statins), is useful for treating all forms of atherosclerosis, as well as the signs, symptoms, or complications associated with it. Atherosclerosis (also known as atherosclerotic vascular disease or ASVD, and coronary artery disease or CAD) is a condition in which the arterial walls thicken as a result of the accumulation of fatty substances such as cholesterol. Atherosclerosis is a chronic disease that can remain asymptomatic for decades. Atherosclerosis is a syndrome that affects arterial blood vessels, and the chronic inflammatory response in the arterial wall is largely due to the accumulation of macrophage leukocytes. This chronic inflammatory response is thought to be promoted by low-density lipoproteins (plasma proteins that transport cholesterol and triglycerides) if lipids and cholesterol are not adequately removed from macrophages by functional high-density lipoproteins (HDL). Atherosclerosis is generally referred to as hardening of the arteries or accumulation of material in the arteries. This is caused by the formation of multiple plaques within the arteries.

[0054] The pathobiology of atherosclerotic lesions is complex, but generally, stable atherosclerotic plaques tend to be asymptomatic and are abundant in the extracellular matrix and smooth muscle cells, while unstable plaques are abundant in macrophages and foam cells. The extracellular matrix (also known as the fibrous capsule) separating the lesion from the ductus arteriosus lumen is usually fragile and prone to rupture. Rupture of the fibrous capsule exposes thrombus-forming materials such as collagen to the bloodstream, ultimately inducing thrombus formation in the lumen. During formation, intraluminal thrombi can completely occlude the artery (e.g., coronary artery occlusion), but more frequently, they can detach and enter the bloodstream, ultimately occluding smaller downstream branches and causing thromboembolism (e.g., carotid artery thrombus formation often leads to stroke). Apart from thromboembolism, chronic enlargement of atherosclerotic lesions can lead to complete occlusion of the lumen. Chronic progression of lesions is often asymptomatic until the luminal narrowing becomes so severe that it leads to insufficient blood supply to downstream tissues, resulting in ischemia.

[0055] These complications of progressive atherosclerosis are chronic, slowly progressing, and cumulative. In some cases, a soft plaque may suddenly rupture, causing the formation of a blood clot, which rapidly slows or stops blood flow and leads to the death of the tissue supplied by the artery (infarction). Coronary thrombosis of the coronary arteries is also a common complication that can lead to myocardial infarction. Blockage of arteries leading to the brain can lead to stroke. In progressive atherosclerotic disease, claudication can occur due to insufficient blood supply to the legs, which can typically be caused by a combination of stenosis and aneurysmal segments narrowed by blood clots.

[0056] Atherosclerosis can affect all arterial trees, but typically, the risk is higher for larger, high-pressure vessels such as the coronary arteries, renal arteries, femoral arteries, cerebral arteries, and carotid arteries.

[0057] Signs, symptoms, and complications of atherosclerosis include, but are not limited to, elevated plasma total cholesterol, VLDL-C, LDL-C, free cholesterol, cholesterol esters, triglycerides, phospholipids, and arterial lesions (e.g., plaques), as discussed above. In some cases, elevated cholesterol (e.g., total cholesterol, free cholesterol, and cholesterol esters) may be found in one or more of the plasma, aortic tissue, and aortic plaques.

[0058] Certain individuals may have a predisposition to atherosclerosis. Therefore, this disclosure relates to a method of administering a CD dimer alone or in combination with one or more additional therapeutic agents (e.g., antihyperlipidemic agents such as statins) to subjects for the prevention of atherosclerosis or its signs, symptoms, or complications. In some embodiments, subjects with a predisposition to atherosclerosis may exhibit one or more of the following characteristics: advanced age, family history of heart disease, biological condition, and high blood cholesterol. In some embodiments, the biological condition includes high levels of low-density lipoprotein cholesterol (LDL-C) in the blood, low levels of high-density lipoprotein cholesterol (HDL-C) in the blood, hypertension, insulin resistance, diabetes, excess body weight, obesity, sleep apnea, contributing lifestyle choices, and / or contributing behavioral habits. In some embodiments, behavioral habits include smoking and / or alcohol consumption. In some embodiments, lifestyle choices include an inactive lifestyle and / or high stress levels.

[0059] In exemplary embodiments, the present disclosure provides administration of the CD dimer of this disclosure in combination with one or more optional additional agonists to a patient suffering from atherosclerosis. The patient may exhibit one or more signs or symptoms of atherosclerosis. Atherosclerosis can be diagnosed based on one or more of the following: Doppler ultrasound, ankle-brachial index, electrocardiogram, stress test, angiography (optionally using cardiac catheterization), computed tomography (CT), magnetic resonance angiography (MRA), or other arterial imaging or blood flow measurement methods.

[0060] In exemplary embodiments, the present disclosure provides administration in combination therapy comprising the CD dimer and one or more additional therapies. These combination therapies for treating atherosclerosis may include the CD dimer of the present disclosure and other therapies for the treatment or prevention of atherosclerosis, such as anticholesterol agents, antihypertensive agents, antiplatelet agents, nutritional supplements, or surgery or behavioral interventions, including, but not limited to, those described below. Further combination therapies may include the CD dimer of the present disclosure and other therapies for the treatment of heart failure, such as one or more or a combination thereof, of aldosterone antagonists, ACE inhibitors, ARBs (angiotensin II receptor blockers), ARNIs (angiotensin receptor neprilysin inhibitors), beta-blockers, vasodilators, calcium channel blockers, digoxin, diuretics, cardiac pump drugs, potassium, magnesium, selective sinoatrial node inhibitors. Combination therapies for treating atrophic age-related macular degeneration (AMD) or Stargardt disease include the CD dimer of this disclosure and another therapy for the treatment of AMD, such as the AREDS nutritional supplement containing LBS-008 (Belite Bio) (a non-retinoid antagonist of retinol-binding protein 4), vitamin C and vitamin E, beta-carotene, zinc, and copper; the AREDS2 nutritional supplement containing vitamin C and vitamin E, zinc, copper, lutein, zeaxanthin, and omega-3 fatty acids; or a combination thereof. Combination therapies for treating Alzheimer's disease include the CD dimer of this disclosure and one or more of the cholinesterase inhibitors (ARICEPT(R), EXELON(R), RAZADYNE(R)) and memantine (NAMENDA(R)) or a combination thereof. Combination therapy for Niemann-Pick disease comprises the CD dimer of this disclosure and one or more of the following: miglustat (ZAVESCA(R)), HPβCD (TRAPPSOL CYCLO, VTS-270), and physiotherapy. This combination therapy may be administered simultaneously, essentially simultaneously, or sequentially in any of these orders.Combination therapy may be administered simultaneously with single formulations or separately, or optionally using a dosing kit or pack containing a combination of each drug, which may be a pre-measured, convenient format providing, for example, one or more single doses of each combined drug. Combination therapy may exhibit synergistic effects, with the combined therapy exceeding the effects of individual monotherapy. Combination therapy generally involves administering effective doses of the C. della bicarbonate dimer and the combined therapy, but combination therapy may yield effective treatment with lower doses of C. della bicarbonate and / or the combined therapy, which may advantageously reduce the side effects associated with the usual (non-combination) doses.

[0061] Combination therapies may include therapies for the treatment or prevention of diseases or conditions associated with atherosclerosis, such as coronary artery disease, angina pectoris, heart attack, cerebrovascular disease, transient ischemic attack, and / or peripheral artery disease. Combination therapies may also include therapies for the treatment or prevention of conditions that may contribute to the formation of atherosclerosis and / or worsening of prognosis, such as hypertension, hypercholesterolemia, hyperglycemia, and diabetes.

[0062] In exemplary embodiments, the CD dimer of the present invention is an anticholesterol agent, for example, a fibrate or statin agent, for example, ADVICOR(R) (niacin sustained-release / lovastatin), ALTOPREV(R) (lovastatin sustained-release), CADUET(R) (amlodipine / atorvastatin combination), CRESTOR(R) (rosuvastatin), JUVISYNC(R) (sitagliptin / simvastatin), LESCOL(R) (fluvastatin), LESCOL It is administered concurrently with XL (fluvastatin sustained-release), LIPITOR(R) (atorvastatin), LIVALO(R) (pitavastatin), MEVACOR(R) (lovastatin), PRAVACHOL(R) (pravastatin), SIMCOR(R) (niacin sustained-release / simvastatin), VYTORIN(R) (ezetimibe / simvastatin), and / or ZOCOR(R) (simvastatin), etc. Anticholesterol agents may be administered in doses effective in preventing or treating hypercholesterolemia.

[0063] In exemplary embodiments, the CD dimer of the present invention is administered concurrently with an antiplatelet drug, such as aspirin.

[0064] In exemplary embodiments, the CD dimer of the present invention is administered concurrently with an antihypertensive drug. Examples of antihypertensive drugs include beta-blockers, angiotensin-converting enzyme (ACE) inhibitors, calcium channel blockers, and / or diuretics.

[0065] In exemplary embodiments, the CD dimer of the present invention is administered concurrently with one or more nutritional supplements, such as alpha-linolenic acid (ALA), barley, beta-sitosterol, black tea, psyllium husk, calcium, cocoa, cod liver oil, coenzyme Q10, fish oil, folic acid, garlic, green tea, niacin, oat bran, omega-3 fatty acids (eicosapentaenoic acid (EPA) and / or docosahexaenoic acid (DHA), etc.), cytostanol, and / or vitamin C.

[0066] Exemplary combination therapies include counseling and / or support for interventions in the patient's behavior and / or lifestyle, such as smoking cessation, exercise, and healthy eating, such as diets low in low-density lipoprotein (LDL) and optionally high in high-density lipoprotein (HDL).

[0067] Exemplary combination therapies include surgical interventions, such as angioplasty, stent placement, or both.

[0068] The methods of the present invention are useful for treating or preventing atherosclerosis in human subjects. In some cases, patients are healthy except for exhibiting atherosclerosis. For example, a patient may not exhibit any other risk factors for cardiovascular, thrombotic, or other diseases or disorders at the time of treatment. However, in other cases, patients are selected based on having been diagnosed with or being at risk of developing a disease or disorder caused by or associated with atherosclerosis. For example, at the time of or before administration of the pharmaceutical composition of the present invention, a patient may have been diagnosed with or been identified as being at risk of developing a cardiovascular disease or disorder, such as coronary artery disease, acute myocardial infarction, asymptomatic carotid atherosclerosis, stroke, peripheral artery occlusive disease, etc. The cardiovascular disease or disorder may, in some cases, be hypercholesterolemia.

[0069] In other cases, at the time of administration of the pharmaceutical composition of the present invention, or prior to administration, the patient may have been diagnosed with or identified as being at risk of developing atherosclerosis.

[0070] In other cases, patients to be treated by the method of the present invention are selected based on one or more factors selected from the group consisting of age (e.g., older than 40, 45, 50, 55, 60, 65, 70, 75, or 80 years of age), race, sex (male or female), exercise habits (e.g., regularly exercises, does not exercise), other pre-existing medical conditions (e.g., type II diabetes, hypertension, etc.), and current medical status (e.g., currently taking statin drugs, e.g., cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin, pravastatin, etc., beta-blockers, niacin, etc.). [Brief explanation of the drawing]

[0071] [Figure 1A] The chemical structures of unsubstituted cyclic oligosaccharides composed of 6(αCD), 7(βCD), or 8(γCD) sugar rings (from left to right) are shown. All sugar rings of all CDs are D-glucose molecules. [Figure 1B] The chemical structures of HPβCD C2DS4, C3DS4, and C6DS4 are shown. [Figure 1C] The chemical structures of randomly substituted MeβCD DS7 are shown. [Figure 1D] The chemical structures of randomly substituted SBβCD DS4 (free acid form) are shown. [Figure 1E] The chemical structures of randomly substituted SUCCβCD DS4 (free acid form) are shown. [Figure 1F] The chemical structures of randomly substituted QAβCD DS4 are shown. [Figure 1G] Dimer structure. The C2-C2 βCD dimer of formula I, linked via a secondary plane using a triazole linker, is shown. [Figure 1H] Dimer structure. It shows the C3-C2 βCD dimer of formula II, linked through a secondary plane using a triazole linker. [Figure 1I] Dimer structure. It shows the C3-C3 βCD dimer of formula III, linked through a secondary plane using a triazole linker. [Figure 1J] Dimerized structure. Shows a hydroxypropyl βCD dimer with secondary planes linked via a variable linker. [Figure 1K] Dimer structure. Shows a methyl βCD dimer linked through a secondary plane using a variable linker. [Figure 2A] Examples of possible substituents are shown. [Figure 2B] Examples of possible linkers, denoted as LAB-A'-L', are shown, where n, n1, n2 = 1, 2, 3, 4, or 5 carbon atoms, and L or L' = L1, L1', L2, or L2'. [Figure 3A] A schematic diagram of a βCD with a "frustum of a cone" shape having a primary (1°) plane and a secondary (2°) plane is shown. [Figure 3B] The structure of 7KC oxysterol, which has a "head group" and a "tail group," is shown. [Figure 3C] The diagram shows the angle measurement between the βCD O4 plane and the ligand axis. [Figure 3D] The diagrams show the βCD monomer-ligand complex in the "up" and "down" directions. While 7KC is used as an example, cholesterol differs only in the carbonyl group at position 7; the same applies to cholesterol. [Figure 3E] The following are presented for unmodified (DS0) monomeric βCD in the GROMOS force field: the distance between the center of gravity of all O4 oxygen atoms and the center of gravity of the ligand in the vertical direction of the ligand; the angle between the vector perpendicular to the plane formed by the O4 atoms of the CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between the CD and the ligand. [Figure 3F] The following are presented for monomeric HPβCD DS5: the distance between the center of gravity of all O4 oxygen atoms and the center of gravity of the ligand in the vertical direction of the ligand under the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4A]The following is presented for the butyl-linked HPβCD DS5 dimer, showing the distance between the centroids of all O4 oxygen atoms in the ligand's vertical direction and the centroid of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4B] The following are presented for the triazole-linked HPβCD DS4 dimer: the distance between the center of gravity of all O4 oxygen atoms in the ligand and the center of gravity of the ligand in the GROMOS force field, in the vertical direction of the ligand; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4C] The following is presented for the butyl-linked MeβCD DS4 dimer, showing the distance between the centroids of all O4 oxygen atoms in the vertical direction of the ligand and the centroid of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4D] The following are presented for the triazole-linked MeβCD DS4 dimer: the distance between the center of gravity of all O4 oxygen atoms in the ligand's vertical direction and the center of gravity of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4E] The following is presented for the butyl-linked SBβCD DS4 dimer: the distance between the center of gravity of all O4 oxygen atoms in the ligand's vertical direction and the center of gravity of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4F]The following are presented for the triazole-linked SBβCD DS4 dimer: the distance between the center of gravity of all O4 oxygen atoms in the ligand's vertical direction and the center of gravity of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4G] The following is presented for the butyl-linked QAβCD DS4 dimer: the distance between the centroids of all O4 oxygen atoms in the ligand's vertical direction and the centroid of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 4H] The following are presented for the triazole-linked QAβCD DS4 dimer: the distance between the center of gravity of all O4 oxygen atoms in the ligand's vertical direction and the center of gravity of the ligand in the GROMOS force field; the angle between the vector perpendicular to the plane formed by the O4 atoms of CD and the principal axis of the ligand; and the MD analysis of the Lennard-Jones and Coulomb energies of the interaction between CD and ligand. [Figure 5A] This shows cholesterol solubilization by various DS HP and MeβCD monomers. [Figure 5B] This shows the solubilization of 7KC by various DS compounds, including HPβCD DS5 and MEβCD monomers. [Figure 5C] This demonstrates the solubilization of cholesterol by various DS-QA, maltosyl, carboxymethyl, succinyl, SBβCD, and HPγCD monomers. [Figure 5D] The solubilization of 7KC by various DS monomers including QA, maltosyl, carboxymethyl, succinylation, SBβCD, and HPγCD is shown. In some examples in the figure, βCD is labeled BCD and γCD is labeled GCD. [Figure 5E] This shows the solubilization of cholesterol and 7KC by various DS-derived HPβCD butyl-linked dimers compared to HPβCD monomers. [Figure 5F]This shows the solubilization of cholesterol and 7KC by HPβCD triazole-linked dimers of various DS compared to HPβCD monomers. [Figure 5G] This shows the solubilization of cholesterol and 7KC by HPβCD DS3 butyl-linked dimers compared to HPβCD monomers. [Figure 5H] This shows the solubilization of cholesterol and 7KC by the HPβCD DS3 triazole-linked dimer compared to the HPβCD monomer. [Figure 5I] This demonstrates the solubilization of cholesterol and 7KC by MeβCD DS3 triazole linkage and HPβCD DS3 triazole linkage dimer. [Figure 5J] This demonstrates the solubilization of cholesterol and 7KC by various DS triazole-linked dimers, including βCD, QAβCD, SBβCD, and SUCCβCD. [Figure 5K] This shows a scatter plot comparing the in vitro performance of βCD monomers and dimers, specifically the 7KC EC50 and in vitro 7KC specificity scores. A low 7KC EC50 indicates that CD has a strong affinity for 7KC. A higher 7KC specificity score indicates that 7KC is solubilized better by CD than cholesterol. Triangles represent βCD butyl-linked dimers, squares represent triazole-linked dimers, and circles represent βCD monomers. [Figure 5L] This bar graph shows a comparison between 7KC EC50 measured by a turbidity assay and the in vitro 7KC specificity scores (black bars) of various substituted βCD monomers across various DSs. [Figure 5M] This bar graph compares the 7KC EC50 measured by a turbidity assay with the in vitro 7KC specificity scores of butyl-linked (diagonal stripes) and triazole-linked (black bars) substituted βCD dimers. [Figure 5N] This shows the percentage of post-treatment red blood cell (RBC) hemolysis under different concentrations and substitutions of triazole-linked and butyl-linked βCD dimers in different DSs. [Figure 5O]This shows the cholesterol concentration (mg / dl) in serum after incubation with whole blood at various concentrations of HPβCD-butyl-DS8 dimers (0-2.5 mM). [Figure 5P] This shows the outflow of 7KC from red blood cells into plasma after treatment with a dimer (0-2.5 mM) of HPβCD-butyl-DS8. [Figure 5Q] This shows the outflow of 7KC from red blood cells into plasma after treatment with HPβCD monomers and DS approximately 5 (0-2.5 mM). [Figure 6A] Schematic cross-sectional diagrams of the spatial interaction between the HPβCD-butyl-DS8 dimer and cholesterol in the CD host-guest complex, as determined by HSQC and ROESY NMR, are shown. In the model on the left, cholesterol is contained in the secondary planes of both HPβCD units, while in the model on the right, the tail of cholesterol is partially contained in the secondary plane of one HPβCD unit. [Figure 6B] A schematic cross-sectional view of the spatial interactions between the HPβCD-butyl-DS8 dimer and 7KC in the CD host-guest complex, as determined by HSQC and ROESY NMR, is shown. [Figure 6C] Schematic cross-sectional diagrams of the spatial interaction between the HPβCD-triazole-DS3 dimer and cholesterol in the CD host-guest complex, as determined by HSQC and ROESY NMR, are shown. In the model on the left, cholesterol is contained in the secondary planes of both HPβCD units, while in the model on the right, the tail of cholesterol is partially contained in the secondary plane of one HPβCD unit. [Figure 6D] A schematic cross-sectional view of the spatial interaction between the HPβCD-triazole-DS3 dimer and 7KC in the CD host-guest complex, as determined by HSQC and ROESY NMR, is shown. [Figure 7A] This document presents a synthetic strategy for a hydroxypropylated dimer linked using a single linker unit based on 1,4-dibromobutane (resulting in the butyl-linked HPβCD dimer, HP-(βCD-butyl-βCD)). [Figure 7B]The MALDI spectrum of the HP-(βCD-butyl-βCD) dimer is shown. [Figure 7C] The structure of one possible isomer of the HP-(βCD-butyl-βCD) dimer is shown with atomic numbers. [Figure 7D] The 1H-NMR spectrum of HP(βCD-BUT-βCD)(D2O, 298K) is shown along with signal labeling and DS calculations. [Figure 7E] The DEPT-edited HSQC spectrum of HP(βCD-BUT-βCD) is shown, with all members assigned (D2O, 298K). It has been converted to black and white. [Figure 7F] This document presents a synthetic strategy for a 2-hydroxypropylated dimer linked to a single linker unit based on 3-azido-1-bromopropane (resulting in the triazole-linked HPβCD dimer, HP-(βCD-triazole-βCD)). [Figure 7G] The MALDI spectrum of the HP-(βCD-triazole-βCD) dimer is shown. [Figure 7H] The structure of one possible isomer of the HP-(βCD-triazole-βCD) dimer is shown with atomic numbers. [Figure 7I] The 1H-NMR spectrum of HP(βCD-triazole-βCD)(D2O, 298K) is shown along with the DS calculation. [Figure 7J] The DEPT-edited HSQC spectrum of HP (βCD-triazole-βCD) is shown, with the linker assigned (D2O, 298K). The image has been converted to black and white. [Figure 7K] The synthesis scheme for the Me-(βCD-triazole-βCD) dimer is shown. [Figure 7L] The MALDI spectrum of the Me-(βCD-triazole-βCD) dimer is shown. [Figure 7M] The structure of one possible isomer of the Me-(βCD-triazole-βCD) dimer is shown with atomic numbers. [Figure 7N]The 1H-NMR spectrum of Me(βCD-triazole-βCD)(D2O, 298K) is shown along with the signal label. [Figure 7O] The COSY-NMR spectrum of the Me-(βCD-triazole-βCD) dimer is shown, and some of its properties have been assigned. [Figure 7P] The DEPT-edited HSQC spectra of the Me-(βCD-triazole-βCD) dimer are shown, all of which have been assigned. The images have been converted to black and white. [Figure 7Q] The synthesis scheme for SB-(βCD-triazole-βCD) dimer is shown. [Figure 7R] The MALDI spectrum of the SB-(βCD-triazole-βCD) dimer low DS is shown. [Figure 7S] One of the possible isomers (sodium salt form) of the SB-(βCD-triazole-βCD) dimer is shown, with atomic numbers assigned to each atom. [Figure 7T] The 1H-NMR spectrum of SB(βCD-triazole-βCD) low DS(D2O, 298K) is shown along with the signal label. [Figure 7U] The COSY spectrum of the SB(βCD-triazole-βCD) dimer low DS is shown and partially assigned (D2O, 298K). [Figure 7V] The DEPT-edited HSQC spectra of the SB-(βCD-triazole-βCD) dimer low DS are shown, all assigned (D2O, 298K). The images have been converted to black and white. [Figure 7W] The MALDI spectrum of SB-(βCD-triazole-βCD) with high DS is shown. [Figure 7X] The 1H-NMR spectrum of SB (βCD-triazole-βCD) high DS (D2O, 298K) is shown. [Figure 7Y] The COSY spectrum of the SB(βCD-triazole-βCD) dimer at high DS is shown and partially assigned (D2O, 298K). [Figure 7Z]The DEPT-edited HSQC spectra of the SB(βCD-triazole-βCD) dimer high DS are shown, all assigned (D2O, 298K). The images have been converted to black and white. [Figure 7AA] The synthesis scheme for the QA-(βCD-triazole-βCD) dimer is shown. [Figure 7AB] The MALDI spectrum of the QA-(βCD-triazole-βCD) dimer is shown. [Figure 7AC] The structure of one possible isomer (DS4) of the QA-(βCD-triazole-βCD) dimer is shown, with atoms numbered. [Figure 7AD] The 1H-NMR spectrum of QA(βCD-triazole-βCD)(D2O, 298K) is shown along with signal labeling and DS calculations. [Figure 7AE] The COSY spectrum of the QA(βCD-triazole-βCD) dimer is shown, and a portion of it has been assigned (D2O, 298K). [Figure 7AF] The DEPT-edited HSQC spectra of the QA(βCD-triazole-βCD) dimer are shown, all assigned (D2O, 298K). The images have been converted to black and white. [Figure 7AG] The synthesis scheme for SUCC-(βCD-triazole-βCD) dimer is shown. [Figure 7AH] The MALDI spectrum of the SUCC-(βCD-triazole-βCD) dimer is shown. [Figure 7AI] The structure (free acid form) of one possible isomer (DS4) of the SUCC-(βCD-triazole-βCD) dimer is shown with atomic numbers. [Figure 7AJ] The 1H-NMR spectrum of SUCC (βCD-triazole-βCD) (D2O, 298K) is shown along with the signal label. [Figure 7AK] The COSY spectrum of the SUCC(βCD-triazole-βCD) dimer is shown, and a portion of it has been assigned (D2O, 298K). [Figure 7AL]The DEPT-edited HSQC spectra of SUCC(βCD-triazole-βCD) dimers are shown, all assigned (D2O, 298K). The images have been converted to black and white. [Figure 8A] The chemical structures of βCD and β'CD are shown as structures A-Xa and A-Xb, respectively, and they combine to form a homodimer by covalent bonding at the L1, L2, L1', and L2' positions on the secondary surface of CD. [Figure 8B] The chemical structure of a homodimer consisting of two βCDs covalently bonded at the L1 and L1' positions on the secondary plane of each CD using a common linker AB-A' is shown. [Figure 8C] The chemical structure of a homodimer consisting of two βCDs covalently bonded at the L1 and L2' positions on the secondary plane of each CD using a common linker AB-A' is shown. [Figure 8D] The chemical structure of a homodimer consisting of two βCDs covalently bonded at the L2 and L2' positions on the secondary plane of each CD using a common linker AB-A' is shown. [Figure 9A] The schematic structure of the dimerized βCD is shown using a common linker represented by AB-A', where AB-A' is as defined herein. Both monomers are randomly substituted with the same functional group on the primary and secondary faces. [Figure 9B] The schematic structure of the βCD homodimer of 2-hydroxypropyl-substituted DS6 is shown using a common AB-A' linker, where AB-A' is as defined herein. [Figure 9C] The schematic structure of a methyl-substituted βCD homodimer is shown using a common AB-A' linker, where AB-A' is defined as herein. [Figure 9D] The schematic structure of the triazole-linked βCD homodimer, where the butyl moiety of the primary DS6 is substituted as evaluated by MD simulation, is shown. [Figure 9E] The schematic structure of the triazole-linked βCD homodimer, in which the (2-hydroxypropyl) moiety of the primary DS6 is substituted as evaluated by MD simulation, is shown. [Figure 10A]The complete chemical structure of one possible isomer of the triazole-linked βCD homodimer, which is substituted with the butyl moiety of the primary DS6 as evaluated by MD simulation, is shown. [Figure 10B] The complete chemical structure of one possible isomer of the triazole-linked βCD homodimer, which is substituted with the (2-hydroxypropyl) moiety of the primary DS6 as evaluated by MD simulation, is shown. [Figure 11A] The results of MD simulations of triazole-linked βCD homodimers substituted with the butyl moiety of primary DS6, which forms a complex with 7KC or cholesterol in both directions, are shown. [Figure 11B] The results of MD simulations of triazole-linked βCD homodimers, in which the (2-hydroxypropyl) moiety of the primary DS6 that forms a complex with 7KC or cholesterol in both directions, are shown. [Figure 12A] The synthesis route for butyl-linked αCD homodimers is shown. [Figure 12B] The synthesis pathway for triazole-linked αCD homodimers is shown. [Figure 13A] The chemical structures of αCD and βCD, structure B-Xa and structure B-Xb, are shown, respectively. These structures combine to form a heterodimer by covalent bonding at the L1, L2, L1', and L2' positions on the secondary surface of CD. [Figure 13B] This shows the "upward" direction of the complex of heterodimer CD and 7KC. [Figure 13C] This shows the "downward" direction of the complex of heterodimer CD and 7KC. [Figure 13D] The chemical structure of the heterodimer consisting of αCD and βCD covalently bonded at the L1 and L1' positions on the secondary plane of each CD using a common linker AB-A' is shown, where AB-A' is as defined herein. [Figure 13E] The chemical structure of the heterodimer consisting of αCD and βCD covalently bonded at the L1 and L2' positions on the secondary plane of each CD using a common linker AB-A' is shown, where AB-A' is as defined herein. [Figure 13F]The chemical structure of the heterodimer consisting of αCD and βCD covalently bonded at the L2 and L2' positions on the secondary plane of each CD using a common linker AB-A' is shown, where AB-A' is as defined herein. [Figure 13G] The schematic structure of an αCD-βCD' heterodimer having a common linker represented by AB-A' is shown, where AB-A' is as defined herein. Both monomers are randomly substituted with the same functional group on the primary and secondary faces. [Figure 13H] The schematic structure of the unmodified αCD-unmodified βCD triazole-linked heterodimer, as evaluated by MD simulation, is shown. [Figure 13I] The schematic structure of the HPDS2αCD-HPDS2βCD triazole-linked heterodimer, as evaluated by MD simulation, is shown. [Figure 13J] The schematic structure of the SBDS2αCD-SBDS2βCD triazole-linked heterodimer, as evaluated by MD simulation, is shown. [Figure 14A] The results of MD simulations of undenatured αCD-undenatured βCD heterodimers that formed complexes with 7KC or cholesterol in both directions are shown. [Figure 14B] The results of MD simulations of HPDS2αCD-HPDS2βCD heterodimers that form complexes with 7KC or cholesterol in both directions are shown. [Figure 14C] The results of MD simulations of SBDS2αCD-SBDS2βCD heterodimers that form complexes with 7KC or cholesterol in both directions are shown. [Figure 15A] The results of the in vitro % turbidity assay obtained from the turbidity assay for the solubilization of 7KC or cholesterol using undenatured αCD, HPβCD DS5, and a 1:1 molar mixed monomer solution of undenatured αCD and HPβCD DS5 at 350 nm are shown. [Figure 15B]The results of the in vitro % turbidity assay obtained from the turbidity assay for the solubilization of 7KC or cholesterol with HP-αCD, HPβCD DS5, and a 1:1 molar mixed monomer solution of HP-αCD and HPβCD DS5 at 350 nm are shown. [Figure 16A] This document describes a synthetic route for producing butyl-linked βCD-αCD heterodimers. [Figure 16B] This document describes the synthetic route for producing triazole-linked βCD-αCD heterodimers. [Figure 16C] This document describes a synthetic route for producing sulfobutylated triazole-linked βCD-αCD heterodimers. [Figure 17A] The two βCD chemical structures, structure C-Xa and structure C-Xb, are shown, and they combine to form an asymmetric dimer covalently bonded at the L1, L2, L1', and L2' positions on the secondary surface of the CD. [Figure 17B] The chemical structure of the asymmetric dimer consisting of βCD covalently bonded at the L1 and L1' positions on the secondary plane of each CD using a common linker AB-A' is shown, where AB-A' is as defined herein. [Figure 17C] The chemical structure of an asymmetric dimer consisting of two βCDs covalently bonded at the L1 and L2' positions on the secondary plane of each CD using a common linker AB-A' is shown, where AB-A' is as defined herein. [Figure 17D] The chemical structure of an asymmetric dimer consisting of two βCDs covalently bonded at the L2 and L2' positions on the secondary plane of each CD using a common linker AB-A' is shown, where AB-A' is as defined herein. [Figure 18A] The schematic structure of the unmodified βCD-HPβCD(DS3-random substitution) asymmetric dimer, as evaluated by MD simulation, is shown. [Figure 18B] The schematic structure of the unmodified βCD-C6HPβCD(DS3) asymmetric dimer, as evaluated by MD simulation, is shown. [Figure 18C]The schematic structure of the unmodified βCD-C6HPβCD(DS7) asymmetric dimer, as evaluated by MD simulation, is shown. [Figure 19A] The results of MD simulations of undenatured βCD-HPβCD(DS3-random substitution) asymmetric dimers that formed complexes with 7KC or cholesterol in both directions are shown. [Figure 19B] The results of MD simulations of undenatured βCD-C6HPβCD(DS3) asymmetric dimers that formed complexes with 7KC or cholesterol in both directions are shown. [Figure 19C] The results of MD simulations of undenatured βCD-C6HPβCD(DS7) asymmetric dimers that formed complexes with 7KC or cholesterol in both directions are shown. [Figure 20A] The first two steps of the synthetic pathway for producing triazole-linked HPβCD-βCD asymmetric dimers are shown. [Figure 20B] The third step of the synthetic route for producing the triazole-linked HPβCD-βCD asymmetric dimer is shown. DS3 is shown. [Figure 20C] The preparation of the azido linker (3-azido-1-bromo-propane) (Step 1A) and the protected 2-hydroxypropylating agent (Step 1B) is shown. [Figure 20D] This shows a portion of the structure of tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD, one of the βCD monomers. [Figure 20E] The completed structures of tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD and 2-O-mono(3-azidopropyl)-βCD are shown, respectively. [Figure 20F] This shows a cycloaddition reaction that produces a triazole-linked HPβCD-βCD asymmetric dimer. DS3 is shown. [Figure 21A] The preparation of the azido linker (3-azido-1-bromo-propane) and its protected version, the 2-hydroxypropylating agent, is shown. [Figure 21B]Step 2: The structures of the two βCD monomers, 2-O-mono(3-azidopropyl)-βCD and the asymmetric monomer per-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD, are shown. [Figure 21C] Step 2 (continued): The structures of the two βCD monomers, 2-O-mono(3-azidopropyl)-βCD and the asymmetric monomer per-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD are shown. [Figure 21D] This shows a cycloaddition reaction that produces the C6HPβCD-triazole-βCD DS7 asymmetric dimer. [Figure 22A] The 1H-NMR spectrum shows (2-hydroxypropyl)-2-O-monopropargyl-βCD at D2O, 298K. [Figure 22B] The 1H-NMR spectrum of (2-hydroxypropyl)-2-O-monopropargyl-βCD at D2O, 298K is shown along with the DS calculation. [Figure 22C] The ¹H-NMR spectrum shows (2-hydroxypropyl)-2-O-monopropargyl-βCD at D2O, 298K, and is partially assigned to this molecule. [Figure 22D] DEPT-edited HSQC spectrum (partially attributed), showing (2-hydroxypropyl)-2-O-monopropargyl-βCD at D2O, 298K. [Figure 23A] The TLC of per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD is shown. [Figure 23B] The MALDI spectrum of per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD is shown. [Figure 23C] The 1H NMR spectrum of per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD is shown. [Figure 23D] The enlarged 1H NMR spectrum of the CD nucleus of per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD (similar to 23C) is shown. [Figure 23E] The 13C NMR spectrum (CDCl3, 298K) of per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD is shown. [Figure 23F] The IR spectrum (CDCl3, 298K) of per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD is shown. [Modes for carrying out the invention]

[0072] I. Definition

[0073] Unless otherwise specified, the following terms used in this specification and in this application, including the claims, have the definitions given herein.

[0074] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural references unless the context makes it readily apparent.

[0075] CD(CD) refers to a cyclic oligosaccharide composed of sugar rings 6(αCD), 7(βCD), or 8(γCD).

[0076] Hydroxypropyl (HP)-substituted CD (CD). As used herein, the terms “hydroxypropyl-substituted CD,” “HP-substituted CD,” “HPβCD,” or “HPαCD” refer to CDs linked to at least one 2-hydroxypropyl group, i.e., -CH2-CH(OH)-CH3. Typically, the HP group is linked to oxygen atoms linked to the C2, C3, and / or C6 carbons of the CD (most commonly, these bonding sites are mixed).

[0077] Sulfobutyl (SB) beta-CD, abbreviated as SBβCD, SBBCD, SB-BCD, SB-βCD, and similar terms, refers to beta-CD substituted with one or more sulfobutyl groups, namely -CH2-CH2-CH2-CH2-SO3H or -CH2-CH2-CH2-CH2-SO3Na, or other salts thereof, which are typically bonded to oxygen atoms bonded to the C2, C3, and / or C6 carbons of the CD (most commonly, these bond sites are mixed).

[0078] QAβCD, QABCD, QA-BCD, QA-βCD, and similar abbreviations, "quaternary ammonium (QA) beta-CD," refer to beta-CD substituted with one or more substituted or unsubstituted quaternary ammonium groups. One substituted quaternary ammonium salt may have the substituted structure trimethylammoniumpropyl, preferably 2-hydroxytrimethylaminopropyl-, i.e., -CH2-CH(OH)-CH2-N + It is (CH3)3. Quaternary ammonium salts are typically bonded to oxygen atoms bonded to the C2, C3, and / or C6 carbons of C2 and C3 (most commonly, these bonding sites are mixed).

[0079] Methylated (Me)beta-CD, abbreviated as MeβCD, MeBCD, Me-BCD, Me-βCD, and similar terms, refers to a beta-CD substituted with one or more methyl groups, i.e., -CH3 groups, which are typically linked to oxygen atoms linked to the C2, C3, and / or C6 carbons of the CD (most commonly, these linkage sites are mixed).

[0080] Carboxymethylated (CM) beta-CD, abbreviated as CMβCD, CMBCD, CM-BCD, CM-βCD, and similar terms, refers to beta-CD substituted with one or more carboxymethyl groups, such as -CH2-CO2H or -CH2-CO2Na, or other salts thereof, which are typically linked to oxygen atoms linked to the C2, C3, and / or C6 carbons of the CD (most commonly, these linkage sites are mixed).

[0081] Succinylated (SUCC) beta-CDs, abbreviated as SUCCβCD, SUCCBD, SUCC-BCD, SUCC-βCD, and similar terms, refer to beta-CDs substituted with one or more succinyl groups, preferably, for example, -CO-CH2-CH2-COOH or -CO-CH2-CH2-COONa, or other salts thereof, which are typically linked to oxygen atoms linked to the C2, C3, and / or C6 carbons of the CD (most commonly, these linkage sites are mixed).

[0082] "C2", "C3", and "C6" refer to carbon positions of glucose subunits having linkers between hydroxyl functional groups or CD monomers, which can be substituted with different substituents (e.g., methyl, hydroxypropyl, sulfobutyl, succinyl, carboxymethyl, and quaternary ammonium functional groups).

[0083] The large surface (secondary surface) refers to the CD monomer side of the glucose subunit, specifically the side containing the hydroxyl group from the C2 and C3 carbons.

[0084] The primary facet (small facet) refers to the CD monomer side of the glucose subunit that contains the hydroxyl group, starting from the C6 carbon.

[0085] The head group refers to the cyclic region of a sterol structure such as cholesterol or 7KC. See Figure 3B.

[0086] The tail group refers to the alkyl region of a sterol structure such as cholesterol or 7KC. See Figure 3B.

[0087] A linker is synonymous with a linking group and is defined as a chemical unit often represented as [AB-A'] that connects to the CD in a CD dimer. Exemplary linkers can connect via L1, L1', L2, or L2' bonded to the above carbon, which may be oxygen or a bond, for example, via the C2 or C3 carbon of each CD subunit.

[0088] Linker length. As used herein, linker length or synonymous “linker length” refers to the number of atoms in the linker in the shortest path through the linker connecting two CD subunits of a CD dimer. In many embodiments, the linker length is the shortest chain of atoms between the terminal atom of A that connects to the CD and the terminal atom of A' that connects to the other CD, and such chain of atoms passes only through atoms of A, B, and A', i.e., referring to structures AX, BX, or CX, the linker length does not include counting atoms of L1, L2, or L3, through which A and A' connect to their respective CD subunits.

[0089] A head-to-head CD dimer refers to a CD dimer in which two CD monomers are linked via the large (secondary) faces of the CD, typically linked via the C2 and / or C3 carbon atoms of each CD monomer.

[0090] A tail-to-tail CD dimer refers to a CD dimer in which two CD monomers are bonded to the minor facet (primary facet) of a CD molecule, typically via the C6 carbon atom of each CD monomer.

[0091] A head-to-tail CD dimer refers to a CD dimer in which two CD monomers are bonded to opposite ends; that is, one monomer is typically bonded via a C6 carbon atom from the minor facet (primary facet), and the other is typically bonded via a C2 and / or C3 carbon atom from the major facet (secondary facet).

[0092] Degree of substitution (DS). As used herein, degree of substitution (DS) represents the amount of substituents attached to a CD monomer or dimer. In general, DS refers to the total number of substitutions at all positions, e.g., positions attached to all C2, C3, and C6 carbons in the CD monomer or dimer (i.e., the number of positions substituted with atoms other than hydrogen). For clarity, in the case of dimers or polymers, DS does not include counting the linker's bond site(s) to each CD subunit, nor does DS include substituents attached only to the linker itself. For example, referring to structure AX (or similarly, structure BX or CX with the necessary modifications) containing structures A-Xa and A-Xb, this term refers to the total number of non-H atoms R1, R1', R2, R2', R3, and R3'. In this example, DS is determined based on the aforementioned R group, regardless of the structure of the corresponding L1, L1', L2, L2', L3, or L3' (e.g., bond, O, S, etc.). The term DS can be used in combination with a specific substituent name to represent the total number of that particular group. The term DS can also be used in combination with the position of substitution (e.g., the C6 position of the D-glucose monomer) to represent the total number of substituents at similar positions around all CD monomers. For example, C6 2-hydroxypropyl DS4 βCD is intended to convey that each CD monomer in the CD dimer has four 2-hydroxypropyl groups attached to the available C6 position. Clearly, the term DS is used to refer to substituents on one or more subunits of CD, which is generally independent of the number of substitutions that may occur elsewhere, such as at the linker that binds the CD dimers. Furthermore, DS can refer to an average value, such as in the case of preparations containing CD molecules with varying numbers of substituents, and therefore can be a non-total value, such as DS approximately 4.2.

[0093] The DS can be measured by known techniques, including mass spectrometry (e.g., matrix-assisted laser desorption / ionization, "MALDI"), or by NMR. MALDI is suitable for CD derivatives having substituents that give a more typical Gaussian distribution of ions in the mass spectrum, as shown for example for methyl, hydroxypropyl, and sulfobutyl substituents (see, for example, Figures 7B, 7G, 7R, 7W, 7AB, 7AH herein and Figures 10G-10I, 10P-10Q, 11C-11G, 11I, 12E, and 12K in U.S. Patent Application No. 16 / 733,945). The average DS when identified by MALDI is calculated by averaging the peak intensities of the signals corresponding to each DS type of the CD in question. In other cases, there may be ion peak patterns that are not usually seen, for example, due to the formation of various adducts, fragments, cleavage products, etc. These problems can potentially be avoided by using other mass spectrometry techniques. Alternatively, NMR can be used to identify peaks corresponding to the protons of the nuclear dimer. First, the DS value can be determined by scaling the measurement so that the peak area corresponds to the known number of such protons in the structure. Then, the signals corresponding to the protons in the substituents are examined and scaled appropriately to obtain the average DS. In simpler cases, obviously decomposed peaks corresponding to the substituent protons are identified and, as described above, have already been scaled. Then, the average number of substituents is obtained by dividing by the number of protons present in that peak. For example, in the case of a hydroxypropyl substituent, a peak identified as corresponding to the 14 protons in the nuclear structure (aromatic region of glucopyranose) is normalized to 14 through identification and signaling. Then, a peak corresponding to the 3 protons of the methyl substituent is identified, and finally, the average number of hydroxypropyl groups per molecule is obtained by dividing the area of ​​that peak by 3. In other examples, substituent peaks and CD nuclear peaks may be very close together or overlap. In this case, the number of contributing protons in the CD nuclear structure is identified and then subtracted from the peak area (the peak area has already been scaled to an integral area of ​​1 per proton), and then the remaining area is divided by the number of contributing protons to obtain the average DS.For example, in the case of methyl substituents (see, for example, Figures 11K–11L shown in U.S. Patent Application No. 16 / 733,945; also see the MeTriDi NMR shown in Figures 7N–7P herein), the cluster of peaks was identified as corresponding to the three methyl hydrogens of the substituent and, additionally, the cluster of 86 protons of the nuclear CD dimer structure. Similar to the example of hydroxypropyl substituents, the peak identified as corresponding to the 14 protons of the nuclear structure (the anomeric region of glucopyranose) was normalized to 14 by identification and signaling; the area of ​​the peak containing methyl hydrogens and nuclear CD hydrogens was determined to be 92.77, and subtracting this signal from the 86 protons of the nuclear CD structure leaves 6.77; after dividing by the three protons of each methyl group, the average DS was estimated to be 2.26. For HP and ME-substituted CDs, divide the integral value by 3; for QA, divide the integral value by 9; for SB, divide the integral value by 2; and for SUCC, divide the integral value by 4. The above calculations are directly applicable to other types of substituents based on the identification of the peak corresponding to the proton in the substituent structure. CD compositions, such as CD dimer compositions (as defined below), may contain mixtures of individual molecules each substituted with a different number of substituents, in which case the DS value is expressed as the average (median) of the number of substitutions. Fractional DS values ​​reflect cases where the median can fall between integer substitutions. Unless otherwise specified, integer DS values ​​indicate CD compositions with a DS number rounded to the nearest integer. For example, DS4 indicates a DS value of at least 3.5 and less than 4.5.

[0094] CD dimer compositions. As used herein, the terms “CD dimer composition” or “CD dimer composition” refer to a mixture of CD dimers, for example, a mixture of CD dimers substituted with varying numbers of the same substituents. Typically, CD dimer compositions are characterized by having a DS identified by a particular substituent. CD dimer compositions can arise from synthetic processes in which substituents are added to CD dimers in a probabilistic manner, largely due to the symmetry of the CD molecule, such that individual CD molecules vary considerably in the number and position of substituents. Furthermore, CD dimer compositions may contain mixtures of individual molecules with different linker bonding sites (e.g., O2 and O2, O2 and O3, O3 and O2, or O3 and O3), or they may have a homogeneous linker bonding site (e.g., O2 and O2 only, O2 and O3 only, O3 and O2 only, or O3 and O3 only). The DS of a CD dimer composition can be identified by NMR and / or mass spectrometry, for example, as described above.

[0095] Terms such as "specifically bind" mean that a molecule, for example, the CD dimer of this disclosure, forms a relatively stable complex with a binding partner, for example, cholesterol (oxysterol, e.g., 7KC), under physiological conditions. Methods for determining whether a molecule specifically binds to a binding partner are well known in the art, and such methods include, for example, equilibrium dialysis and surface plasmon resonance. In exemplary embodiments, the CD dimer of this disclosure is available in concentrations of approximately 5 μM to 100 μM, approximately 10 μM to 90 μM, approximately 20 μM to 80 μM, approximately 30 μM to 70 μM, approximately 40 μM to 60 μM, approximately 0.5 μM to 50 μM, approximately 1 μM to 40 μM, approximately 2 μM to 30 μM, approximately 3 μM to 20 μM, approximately 4 μM to 10 μM, less than approximately 1000 μM, less than approximately 500 μM, less than approximately 300 μM, It binds to cholesterol, oxysterols, or 7KC at KD levels of less than approximately 200 μM, less than approximately 100 μM, less than approximately 90 μM, less than approximately 80 μM, less than approximately 70 μM, less than approximately 60 μM, less than approximately 50 μM, less than approximately 30 μM, less than approximately 20 μM, less than approximately 10 μM, less than approximately 5 μM, less than approximately 4 μM, less than approximately 3 μM, less than approximately 2 μM, less than approximately 1 μM, or less than approximately 0.5 μM.

[0096] Affinity for 7KC higher than affinity for cholesterol. As used herein, the term “affinity for 7KC higher than affinity for cholesterol” means that a compound (e.g., CD) has a higher ability to solubilize 7KC than cholesterol. Higher affinity can also be predicted by molecular docking, by molecular dynamics simulation, or by calorimetry. In exemplary embodiments, a CD dimer has a binding affinity for 7KC that is at least 1.5 times, at least 2 times, at least 3 times, at least 4 times, at least 5 times, at least 8 times, at least 10 times, at least 15 times, at least 20 times, at least 30 times, or at least 50 times stronger than its binding affinity for cholesterol, which can be optionally determined by comparing the concentration at which 50% of 7KC in a suspension is solubilized, for example, using the procedures described herein in the examples. In exemplary embodiments, the CD dimer has a binding affinity to 7KC that is at least 1.1 times, 1.5 times, 2 times, 3 times, 4 times, 5 times, or 10 times stronger than its binding affinity to cholesterol, which can optionally be determined by dividing the computer-calculated or measured binding affinity (KD) to cholesterol by the computer-calculated binding affinity to 7KC.

[0097] A higher affinity for one compound than for another, for example, a higher affinity for 7KC than for cholesterol, can be determined using a "turbidity test" performed with an aqueous suspension containing 3% ethanol, 300 μM sterols, and 1 mM of the CD under test in PBS. This single concentration of CD is used for the purpose of normalizing the test results. To perform the test, the sample is incubated at 37°C for 30 minutes, and then the absorbance at 350 nm is measured, for example, using a spectrophotometer plate reader. Relative turbidity is determined by dividing the turbidity measured in the presence of CD by the baseline turbidity without CD. If the relative turbidity of the 7KC suspension is lower than the relative turbidity of the cholesterol solution, then a given CD has a higher affinity for 7KC than for cholesterol.

[0098] Hydrophobic drugs. As used herein, the term “hydrophobic drug” refers to a drug that does not dissolve in water unless certain surfactants or other solvents are present. Examples of hydrophobic drugs include, but are not limited to, hormones, estrogen, progesterone, and testosterone. The CD dimers of this disclosure may be used as excipients for hydrophobic drugs. Additional examples of hydrophobic drugs include, to name a few, dexamethorphan (DXM), diphenhydramine (DPH), lidocaine (LDC), bendroflumethiazide, acyclovir, levaprazan, curcumin, and testosterone propionate (TP). The CD dimer may be present in an amount sufficient to increase the solubility of the molecule and / or to aid in better drug delivery. The molecular ratio of drug to CD may be 1:1 or greater.

[0099] An amount effective in solubilizing the hydrophobic drug. As used herein, the phrase "an amount effective in solubilizing the hydrophobic drug" typically refers to the concentration of a substance (e.g., one or more CD dimers) capable of solubilizing the hydrophobic drug in an aqueous composition, such as phosphate-buffered saline (PBS) or water. Solubilization can be measured by spectroscopic or other means known in the art. Solubilization may be measured at room temperature, physiological temperature (37°C), or another suitable temperature (e.g., 0°C to 4°C).

[0100] Heterodimer. As used herein, a heterodimer refers to two different CD monomeric forms covalently bonded together using the linker AB-A' (i.e., αCD-AB-A'-βCD).

[0101] Homodimer. As used herein, homodimer refers to two identical CD monomer forms having the same functional group, covalently bonded by a linker such as [AB-A'] (i.e., βCD-[AB-A']-βCD').

[0102] Asymmetric dimer. As used herein, an asymmetric dimer refers to two CD monomers having different combinations of substitutions on each CD monomer, covalently bonded by a linker such as AB-A'. Non-limiting examples of asymmetric dimers include dimers having two subunits, each containing a different number of the same substituents; dimers containing different substituents; dimers having one substituent at one position on one monomer and the same or different substituents at different positions on the other monomer (e.g., the C2 or C3 substituent on one monomer) and the C6 substituent on the other monomer; dimers where one monomer is substituted and the other is unsubstituted; and dimers having a positively charged substituent on one monomer and a negatively charged substituent on the other monomer. Combinations of the above, such as dimers containing different types and different numbers of substituents on each monomer, are also conceivable.

[0103] Molecular dynamics (MD) refers to computer simulation methods that use GROMACS software (e.g., via GROMOS 54a7) to identify intermolecular interactions of CD-sterol complexes.

[0104] "Upward direction" refers to the position of cholesterol and / or 7KC relative to CD, where the sterol head group is related to the minor / primary plane and the tail group is related to the major / secondary plane. In the case of heterodimers, upward direction refers to the position where the sterol head group is in the βCD sister monomer and the tail group is in the αCD sister monomer.

[0105] "Downward" refers to the position of cholesterol and / or 7KC relative to CD, where the tail group of the sterol is related to the minor / primary plane and the head group is related to the major / secondary plane. In the case of heterodimers, downward refers to the position where the head group of the sterol is in the αCD sister monomer and the tail group is in the βCD sister monomer.

[0106] The O4 plane (or O4 axis) refers to the plane formed by the O4 oxygen atoms of the glucose units that make up the CD molecule. According to the standard nomenclature for glucose units, O4 indicates the number of oxygen atoms (4). See Figure 3.

[0107] The term "angle," used in conjunction with the O4 plane, for example, "O4 plane angle," refers to the angle between the O4 plane of one CD monomer and the ligand axis, indicating how well the ligand is contained within the CD cavity. Angle measurements can be useful in determining how well the ligand is shielded from surrounding water molecules; zero or 180 degrees indicates the ligand is perpendicular to the O4 plane of the CD, and therefore the two molecules are most likely to be in a soluble complex, while 90 degrees indicates the ligand is parallel to the CD plane and less likely to form a complex within the cavity. In this case, approximately 30 degrees corresponds to complex formation in the "up" direction, and approximately 150 degrees corresponds to complex formation in the "down" direction.

[0108] In MD simulations, the distance refers to the distance between the center of mass of the sterol and the center of mass of the CD dimer.

[0109] The energy in MD simulations refers to the energy of the interaction between sterols and CD dimers.

[0110] "Alkyl" refers to a straight-chain or branched-chain hydrocarbon portion consisting only of carbon atoms and hydrogen atoms.

[0111] "Lower alkyl" refers to alkyl groups with 1 to 6 carbon atoms, i.e., C3 alkyl groups. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, isobutyl, sec-butyl, tert-butyl, pentyl, n-hexyl, octyl, and dodecyl.

[0112] "Heteroalkyl" refers to a linear or branched hydrocarbon moiety in which at least one carbon atom is substituted by a heteroatom selected from a list consisting of oxygen, nitrogen, or sulfur, or optionally, silicon or phosphorus. Examples include, but are not limited to, alkoxyalkyls, alkoxyalkoxyalkyls, alkylcarbonyloxyalkyls, alkylcarbonyls, alkylsulfonyls, alkylsulfonylalkyls, alkylaminos, alkylsulfanyls, alkylaminoalkyls, aminoalkyls, dialkylaminoalkyls, aminoalkoxys, alkylsulfonylamides, aminocarbonyloxyalkyls, aminosulfonyls, alkylaminosulfonyls, or dialkylaminosulfonyls.

[0113] "Alkenyl" refers to a linear monovalent hydrocarbon radical consisting of 2 to 12 carbon atoms or a branched monovalent hydrocarbon radical consisting of 3 to 12 carbon atoms, containing at least one double bond. Examples of alkenyl groups include, but are not limited to, ethenyl (vinyl, -CH=CH2), 1-propenyl (-CH=CH-CH3), and 2-propenyl (allyl, -CH-CH=CH2) moieties. Examples include, but are not limited to, methoxy, ethoxy, isopropoxy, etc.

[0114] "Alkoxyalkyl" refers to the portion of the formula Ra-O-Rb-, where Ra is alkyl and Rb is alkylene, as defined herein. Examples of alkoxyalkyl groups include, for example, 2-methoxyethyl, 3-methoxypropyl, 1-methyl-2-methoxyethyl, 1-(2-methoxyethyl)-3-methoxypropyl, and 1-(2-methoxyethyl)-3-methoxypropyl.

[0115] "Alkoxyalkoxyalkyl" means a group of the formula -RO-R'-O-R'', where R and R' are alkylenes and R'' is alkyl, as defined herein.

[0116] "Alkylcarbonyloxyalkyl" means a group of the formula -ROC(O)-R', where R is alkylene and R' is alkyl, as defined herein.

[0117] "Alkylcarbonyl" refers to the part of the formula -R'-R'', where R' is -C(=O)- and R'' is alkyl, as defined herein.

[0118] "Alkylsulfonyl" refers to the part of the formula -R'-R'', where R' is -SO2- and R'' is alkyl, as defined herein.

[0119] "Alkylsulfonylalkyl" refers to the part of the formula -R'-R''-R''', where R' is alkyl, R'' is -SO2-, and R''' is alkyl, as defined herein.

[0120] "Alkylamino" means the part of the formula -NR-R', where R is hydrogen or alkyl and R' is alkyl, as defined herein.

[0121] "Aminoalkyl" means the group -R-R', where R' is amino and R is alkylene, as defined herein. Examples of "aminoalkyl" include aminomethyl, aminoethyl, 1-aminopropyl, and 2-aminopropyl.

[0122] "Dialkylaminoalkyl" means the group -R-NR'R'', where R is alkylene and R' and R'' are alkyl, as defined herein. Examples of dialkylaminoalkyls include dimethylaminomethyl, dimethylaminoethyl, dimethylaminopropyl, and N-methyl-N-ethylaminoethyl.

[0123] "Aminoalkoxy" means the group -OR-R', where R' is amino and R is alkylene, as defined herein.

[0124] "Alkylaminoalkyl" refers to the group -R-NHR', where R is alkylene and R' is alkyl. Examples of alkylaminoalkyl groups include methylaminomethyl, methylaminoethyl, methylaminopropyl, and ethylaminoethyl.

[0125] "Alkylsulfanyl" refers to the -SR portion of the formula, where R is alkyl, as defined herein.

[0126] "Alkali metal ions" refers to Group I metals, such as lithium, sodium, potassium, rubidium, or cesium, preferably monovalent ions of sodium or potassium.

[0127] "Alkaline earth metal ions" refers to Group II metals, such as beryllium, magnesium, calcium, strontium, or barium, preferably divalent ions of magnesium or calcium.

[0128] "Alkylsulfonylamide" refers to the part of the formula -NR'SO2-R, where R is alkyl and R' is hydrogen or alkyl.

[0129] "Aminocarbonyloxyalkyl" or "carbamylalkyl" means the group -ROC(=O)-R', where R' is amino and R is alkylene, as defined herein.

[0130] "Aminosulfonyl" means the group -SO2-NR'R'', where R' and R'' are independently hydrogen or alkyl. Thus, as used herein, "aminosulfonyl" encompasses "alkylaminosulfonyl" and "dialkylaminosulfonyl".

[0131] "Alkynylalkoxy" means a group of the formula -OR-R', where R is alkylene and R' is alkynyl, as defined herein.

[0132] "Cycloalkyl" refers to a saturated or partially unsaturated carbocyclic moiety consisting of one or more rings. Examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and their partially unsaturated derivatives.

[0133] "Hypercycloalkyl" refers to a saturated or partially unsaturated carbocyclic moiety consisting of one or more rings in which at least one carbon atom is substituted with a heteroatom selected from the list of oxygen, nitrogen, or sulfur, or optionally with silicon or phosphorus.

[0134] "Aryl" means a cyclic aromatic hydrocarbon moiety consisting of a monocyclic, dicyclic, or tricyclic system, including fused ring systems. The aryl group can be optionally substituted as defined herein. Examples of aryl moieties include, but are not limited to, optionally substituted phenyl, naphthyl, phenanthryl, fluorenyl, indenyl, pentarenyl, azlenyl, oxydiphenyl, biphenyl, methylenediphenyl, aminodiphenyl, diphenylsulfidyl, diphenylsulfonyl, diphenylisopropylidenyl, benzodioxanyl, benzofuranyl, benzodioxylyl, benzopyranyl, benzooxazinyl, benzooxazinyl, benzopiperazinyl, benzopyrrolidinyl, benzomorpholinyl, methylenedioxyphenyl, ethylenedioxyphenyl, and their partially hydrogenated derivatives.

[0135] "Heteroaryl" means a cyclic aromatic moiety having at least one ring, where at least one ring contains at least one heteroatom selected from the list of O, N, or S, and the remaining ring atoms are C. The heteroaryl ring may be optionally substituted as defined herein. Examples of heteroaryl moieties include, but are not limited to, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyrazinyl, thienyl, benzothienyl, thiophenyl, furanil, pyranyl, pyridyl, pyrrolyl, pyrazolyl, pyrimidyl, quinolinyl, isoquinolinyl, benzofuryl, benzothiophenyl, benzothiopyranil, benzimidazolyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzopyranil, indolyl, isoindolyl, triazole, triazinyl, quinoxalinyl, purinyl, quinazolinyl, quinolidinyl, naphthilidinyl, pteridinyl, carbazolyl, azepinyl, diazepinyl, acridinyl, etc., and their partially hydrogenated derivatives.

[0136] "Amine" or "amino" means the group -NR'R'', where R' and R'' are independently hydrogen or alkyl. Thus, as used herein, "amino" encompasses "alkylamino" and "dialkylamino".

[0137] "Alkoxyamine" or "Alkoxyamino" means the group -OR-R', where R' is amino and R is alkylene, as defined herein.

[0138] Multiple linkers refer to multiple, preferably identical, links between two CD monomers that interact with the C2 or C3 hydroxyl group of the glucose subunit of each CD monomer.

[0139] As used in this specification, the terms “halogen,” “halo,” and “halide” refer to any of the following groups: -F, -Cl, -Br, and -I. In certain embodiments, these groups are specifically referred to as fluoro-, chloro-, bromo-, and iodo-.

[0140] Any open valency appearing in carbon, oxygen, sulfur, or nitrogen atoms in the structures described herein indicates the presence of a hydrogen atom.

[0141] Unless otherwise specified, when any of the above groups is described as “substituted” herein, it should be understood that one or more hydrogens of that group are substituted by any group as defined in the Definitions section or elsewhere herein, such groups include alkyl, cycloalkyl, cycloalkylalkyl, heteroalkyl, hydroxyalkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, acylamino, monoalkylamino, dialkylamino, haloalkyl, haloalkoxy, heteroalkyl, -COR (wherein R is hydrogen, alkyl, phenyl, or phenylalanyl Examples include, but are not limited to, -(CR'R'')n-COOR (wherein n is an integer from 0 to 5, R' and R'' are independently hydrogen or alkyl, and R is hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, phenyl, or phenylalkyl), or -(CR'R'')n-CONRaRb (wherein n is an integer from 0 to 5, R' and R'' are independently hydrogen or alkyl, and Ra and Rb are independently hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, phenyl, or phenylalkyl). These groups substituted with hydrogen may be substituted where applicable. However, none of the substituents of a substituent can be further substituted.

[0142] For all chemical groups defined above, it should be understood that all groups, regardless of whether "-yl", "-ylene", or other suffixes are used, have at least a sufficient number of valencies to satisfy any connectivity required by the more general chemical structure. As a non-restrictive example, if variable B is shown as having connectivity to variables A and A', any selection of variable B will have at least two valencies, even if the cited selection suffix ends in "-yl" or another suffix, meaning there are fewer than two valencies available for bonding. Continuing from this non-restrictive example, if B is selected as a heteroaryl group, it should be understood that any selected "heteroaryl" group will have at least two valencies available for bonding with variables A and A'.

[0143] In certain embodiments, it may be useful to describe the two variable groups of a chemical structure as “each pair.” Each pair can also be represented by listing the two variables separated by a slash (e.g., L1 / R1). The term “each pair” should be understood to mean that each choice of variable in each pair follows a subsequent list of choices for each variable. As a non-restrictive example, the statement “Each pair of L1 / R1 is a bond and a hydroxyl group” is defined here to convey that L1 is a bond and R1 is a hydroxyl group.

[0144] In certain embodiments, the term “corresponding” is used to refer to elements that are shown linked to each other in a structural formula. For example, in structures A-Xa, B-Xa, and C-Xa, each R1 is shown linked to each L1, and the L1 and R1 element pairs showing that they are linked are considered to correspond to each other. Similarly, each of R2 and R3 has corresponding L2 and L3 in structures A-Xa, B-Xa, and C-Xa, respectively, and in structures A-Xb, B-Xb, and C-Xb, R1', R2', and R3' have corresponding L1', L2', and L3' to which they are linked, respectively.

[0145] The terms "arylalkyl" and "aralkyl" are synonymous and refer to radical-RaRb, where Ra is an alkylene group and Rb is an aryl group, as defined herein. For example, phenylalkyls such as benzyl, phenylethyl, and 3-(3-chlorophenyl)-2-methylpentyl are examples of arylalkyls.

[0146] "Arylsulfonyl" means a group of the formula -SO2-R, where R is aryl, as defined herein.

[0147] "Aryloxy" refers to a group of the formula -OR, where R is aryl, as defined herein.

[0148] "Aralkyloxy" or "arylalkyloxy" means a group of the formula -OR-R'', where R is alkylene and R' is aryl, as defined herein.

[0149] "Cyanoalkyl" refers to the part of the formula -R'-R'', where R' is an alkylene as defined herein and R'' is cyano or nitrile.

[0150] A "cycloalkenyl" refers to a monovalent unsaturated carbocyclic moiety consisting of a monocyclic or bicyclic ring and having at least one double bond. Cycloalkenyls can be optionally substituted with one or more substituents, each substituent independently being hydroxy, alkyl, alkoxy, halo, haloalkyl, amino, monoalkylamino, or dialkylamino unless otherwise specifically indicated. Examples of cycloalkenyl moieties include, but are not limited to, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, and cycloheptenyl.

[0151] "Cycloalkylalkyl" refers to the part of the formula -R'-R'', where R' is alkylene and R'' is cycloalkyl, as defined herein.

[0152] "Cycloalkylene" refers to a divalent saturated carbocyclic radical consisting of a monocyclic or bicyclic ring. Cycloalkylenes can be optionally substituted with one or more substituents, each substituent independently being hydroxy, alkyl, alkoxy, halo, haloalkyl, amino, monoalkylamino, or dialkylamino unless otherwise specifically indicated.

[0153] "Cycloalkylalkylene" refers to the part of the formula -R'-R''-, where R' is alkylene and R'' is cycloalkylene, as defined herein.

[0154] "Heteroarylalkyl" or "heteroaralkyl" means a group of the formula -R-R', where R is alkylene and R' is heteroaryl, as defined herein.

[0155] "Heteroarylsulfonyl" refers to a group of the formula -SO2-R, where R is heteroaryl, as defined herein.

[0156] "Heteroaryloxy" means a group of the formula -OR, where R is heteroaryl, as defined herein.

[0157] "Heteroaralkyloxy" means a group of the formula -OR-R'', where R is alkylene and R' is heteroaryl, as defined herein.

[0158] "Heterocycloalkylene" means a cycloalkylene as defined herein, in which one or more carbon atoms are substituted with heteroatoms selected from N, O, or S.

[0159] "Heterocyclylalkoxy" means a group of the formula -OR-R', where R is alkylene and R' is heterocyclyl, as defined herein.

[0160] "Haloalkyl" means an alkyl group as defined herein, in which one or more hydrogen atoms are substituted with the same or different halogens. In some embodiments, the haloalkyl is a fluoroalkyl group. In some embodiments, the haloalkyl is a perfluoroalkyl group. Examples of haloalkyl groups include -CH2Cl, -CH2CF3, -CH2CCl3, and perfluoroalkyl groups (e.g., -CF3).

[0161] "Haloalkoxy" refers to the part of the formula -OR, where R is a haloalkyl part, as defined herein. In some embodiments, the haloalkoxy is a fluoroalkoxy. In some embodiments, the haloalkoxyl is a perfluoroalkoxy. An example of a haloalkoxy is difluoromethoxy.

[0162] "Heterocycloamino" refers to a saturated ring in which at least one ring atom is N, NH, or N-alkyl, and the remaining ring atoms form alkylene groups.

[0163] "Heterocyclyl" means a monovalent saturated moiety consisting of 1 to 3 rings and incorporating 1, 2, 3 or 4 heteroatoms (selected from nitrogen, oxygen or sulfur). The heterocyclyl ring may optionally be substituted as defined herein. Examples of heterocyclyl moieties include optionally substituted piperidinyl, piperazinyl, homopiperazinyl, azepinyl, pyrrolidinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, pyridinyl, pyridazinyl, pyrimidinyl, oxazolidinyl, isoxazolidinyl, morpholinyl, thiazolidinyl, isothiazolidinyl, quinuclidinyl, quinolinyl, isoquinolinyl, benzimidazolyl, thiadiazolidinyl, benzothiazolidinyl, benzoazolidinyl, dihydrofuryl, tetrahydrofuryl, dihydropyranyl, tetrahydropyranyl, thiomorpholinyl, thiomorpholinyl sulfoxide, thiomorpholinyl sulfone, dihydroquinolinyl, dihydroisoquinolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl and the like, but are not limited thereto.

[0164] "Heterocyclylalkyl" means a moiety of the formula -R-R', wherein R is alkylene and R' is heterocyclyl, as defined herein.

[0165] "Heterocyclyloxy" means a moiety of the formula -OR, wherein R is heterocyclyl, as defined herein.

[0166] "Heterocyclylalkoxy" means a moiety of the formula -OR-R', wherein R is alkylene and R' is heterocyclyl, as defined herein.

[0167] "Hydroxyalkoxy" means a moiety of the formula -OR, wherein R is hydroxyalkyl, as defined herein.

[0168] "Hydroxyalkylamino" refers to the part of the formula -NR-R', where R is hydrogen or alkyl and R' is hydroxyalkyl, as defined herein.

[0169] "Hydroxyalkylaminoalkyl" means the part of the formula -R-NR'-R'', where R is alkylene, R' is hydrogen or alkyl, and R'' is hydroxyalkyl, as defined herein.

[0170] "Hydroxyalkyl" means a molecule in which the alkyl portion, as defined herein, is substituted with one or more, preferably one, two, or three hydroxyl groups, provided that no single carbon atom has more than one hydroxyl group. Representative examples include, but are not limited to, hydroxymethyl, 2-hydroxyethyl, 2-hydroxypropyl, 3-hydroxypropyl, 1-(hydroxymethyl)-2-methylpropyl, 2-hydroxybutyl, 3-hydroxybutyl, 4-hydroxybutyl, 2,3-dihydroxypropyl, 2-hydroxy-1-hydroxymethylethyl, 2,3-dihydroxybutyl, 3,4-dihydroxybutyl, and 2-(hydroxymethyl)-3-hydroxypropyl.

[0171] "Hydroxycarbonylalkyl" or "carboxyalkyl" means a group of the formula -R-(CO)-OH, where R is alkylene, as defined herein.

[0172] "Hydroxyalkyloxycarbonylalkyl" or "hydroxyalkoxycarbonylalkyl" refers to a group of the formula -RC(O)-OR-OH, where each R is an alkylene, and they may be the same or different.

[0173] "Hydroxyalkyl" means a molecule in which the alkyl portion, as defined herein, is substituted with one or more, preferably one, two, or three hydroxyl groups, provided that no single carbon atom has more than one hydroxyl group. Representative examples include, but are not limited to, hydroxymethyl, 2-hydroxyethyl, 2-hydroxypropyl, 3-hydroxypropyl, 1-(hydroxyl-5-methyl)-2-methylpropyl, 2-hydroxybutyl, 3-hydroxybutyl, 4-hydroxybutyl, 2,3-dihydroxypropyl, 2-hydroxy-1-hydroxymethylethyl, 2,3-dihydroxybutyl, 3,4-dihydroxybutyl, and 2-(hydroxymethyl)-3-hydroxypropyl.

[0174] "Hydroxycycloalkyl" means a cycloalkyl moiety as defined herein, in which one, two, or three hydrogen atoms of a cycloalkyl radical are substituted with hydroxy substituents. Typical examples include, but are not limited to, 2-, 3-, or 4-hydroxycyclohexyl.

[0175] "Urea" refers to a group of the formula -NR'-C(O)-NR''R''', where R, R'', and R''' are independently hydrogen or alkyl.

[0176] "Carbamate" refers to a group of the formula -OC(O)-NR'R'', where R' and R'' are independently hydrogen or alkyl.

[0177] "Carboxylate" refers to the group with the formula -C(O)OH.

[0178] "Sulfonamide" refers to a group of the formula -SO2-NR'R'', where R', R'', and R'' are independently hydrogen or alkyl.

[0179] "Nitro" means -NO2.

[0180] "Cyano" means -CN.

[0181] "Phenoxy" refers to a phenyl ring substituted with at least one -OH group.

[0182] "Acetyl" means -C(=O)-CH3.

[0183] "Cn-m-" is used as a prefix before a functional group, for example, C1-12-alkyl or C5-12-heteroaryl, where "n" and "m" are integer values ​​(i.e., 0, 1, 2, 12). This prefix indicates the number or range of carbon atoms present in the functional group. In the case of cyclic systems, this prefix indicates the number or range of ring atoms, regardless of whether the ring atoms are carbon atoms or heteroatoms. When a functional group consists of a cyclic and acyclic part (i.e., in the case of "arylalkyl," it consists of an aryl part and an alkyl part), this prefix is ​​used to indicate how many carbon atoms and ring atoms are present in total. For example, in the case of arylalkyl, "C7-arylalkyl" can be used to indicate "phenyl-CH2-". In some functional groups, there may be zero carbon atoms present; for example, in C0-aminosulfonyl (i.e., -SO2-NH2, where both possible R groups are hydrogen), "0" indicates the absence of carbon atoms.

[0184] In organic synthesis, the term "leaving group" refers to a group that has traditionally been associated with this term, that is, an atom or group that can be replaced under substitution conditions. Examples of leaving groups include, but are not limited to, halogens, alkanes, or arylenesulfonyloxy groups, such as methanesulfonyloxy, ethanesulfonyloxy, trifluoromethanesulfonyloxy, thiomethyl, benzenesulfonyloxy, tosyloxy, and thienyloxy, as well as dihalophosphinoloxy, ammonium quaternaries, and optionally substituted benzyloxy, isopropyloxy, and acyloxy groups.

[0185] "Modulator" means a molecule that interacts with a target. Examples of interactions include, but are not limited to, agonists, antagonists, etc. as defined herein.

[0186] "Optional" or "optionally" means that the subsequently described event or situation may occur, but does not necessarily occur, and that the description includes cases where the event or situation occurs and cases where the event or situation does not occur.

[0187] "Disease" and "disease state" mean any disease, symptom, syndrome, disorder, or sign.

[0188] "Inert organic solvent" or "inert solvent" means that the solvent is inert under the reaction conditions described in conjunction with the solvent. Examples of such solvents include, for example, benzene, toluene, acetonitrile, tetrahydrofuran, N,N-dimethylformamide, chloroform, methylene chloride or dichloromethane, dichloroethane, diethyl ether, ethyl acetate, acetone, methyl ethyl ketone, methanol, ethanol, propanol, isopropanol, tert-butanol, dioxane, pyridine, etc. Unless otherwise specified, the solvents used in the reactions of the present disclosure are inert solvents.

[0189] "Pharmaceutically acceptable" means generally safe, non-toxic, and useful for the preparation of pharmaceutical compositions that are not undesirable biologically or otherwise, and includes those acceptable not only for human pharmaceutical use but also for veterinary use.

[0190] A "pharmaceutically acceptable salt" of a compound means a pharmaceutically acceptable salt as defined herein that retains the desired pharmacological activity of the parent compound. Examples of such salts include acid addition salts formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; or acid addition salts formed with organic acids, such as acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, glucoheptonic acid, gluconic acid, glutamic acid, glycolic acid, hydroxynaphthoic acid, 2-hydroxyethanesulfonic acid, lactic acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, muconic acid, 2-naphthalenesulfonic acid, propionic acid, salicylic acid, succinic acid, tartaric acid, p-toluenesulfonic acid, trimethylacetic acid, etc.; or salts formed when an acidic proton present in the parent compound is substituted with a metal ion, such as an alkali metal ion, an alkaline earth ion, or an aluminum ion, or when an organic or inorganic base is coordinated. Acceptable organic bases include diethanolamine, ethanolamine, N-methylglucamine, triethanolamine, trimethylamine, and tromethamine. Acceptable inorganic bases include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, and sodium hydroxide. Preferred pharmaceutically acceptable salts are those formed from acetic acid, hydrochloric acid, sulfuric acid, methanesulfonic acid, maleic acid, phosphoric acid, tartaric acid, citric acid, sodium, potassium, calcium, zinc, and magnesium. All references to pharmaceutically acceptable salts include the solvated form (solvate) or crystalline form (polymorph) of the acid addition salt as defined herein. In general, if a particular salt is included in a structure or formula herein, it is possible to substitute it with another pharmaceutically acceptable salt within the scope of this disclosure. For example, in the case of the quaternary ammonium salt of formula VIII, this may include a chloride or another anion or combination of ions, and similarly, in the case of the carboxymethyl sodium salt of formula IX, another cation may substitute for the indicated sodium.

[0191] When used herein, the term “pharmaceutically acceptable carrier” generally refers to a pharmaceutically acceptable composition useful for introducing an active agent into the body, such as a liquid or solid filler, diluent, excipient, manufacturing aid (e.g., lubricant, magnesium talc, calcium stearate or zinc stearate, or stearic acid), or solvent encapsulation material. Each carrier must be “acceptable” in the sense that it is compatible with the other components of the formulation and is not harmful to the patient. Examples of suitable aqueous and non-aqueous carriers usable in the pharmaceutical compositions of the present invention include, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.), vegetable oils (e.g., olive oil), and organic esters for injection (e.g., ethyl oleate), as well as suitable mixtures thereof. Adequate fluidity can be maintained, for example, by the use of a coating material such as lecithin, by maintaining the required particle size in the case of dispersion, and by the use of a surfactant.

[0192] Other examples of materials that can function as pharmaceutically acceptable carriers include: (1) sugars, e.g., lactose, glucose, and sucrose; (2) starches, e.g., corn starch and potato starch; (3) cellulose and its derivatives, e.g., sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, e.g., cocoa butter and suppository wax; (9) oils, e.g., peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; (10) glycols. Examples include (11) propylene glycol, polyols such as glycerin, sorbitol, mannitol, and polyethylene glycol, (12) esters such as ethyl oleate and ethyl laurate, (13) agar, (14) buffers such as magnesium hydroxide and aluminum hydroxide, (15) alginic acid, (16) pyrogen-free water, (17) isotonic saline, (18) Ringer's solution, (19) ethyl alcohol, (20) pH buffer, (21) polyesters, polycarbonates, and / or polyanhydrides, and (22) other non-toxic and suitable substances used in pharmaceutical formulations.

[0193] Various auxiliary agents such as wetting agents, emulsifiers, lubricants (e.g., sodium lauryl sulfate and magnesium stearate), colorants, release agents, coating agents, sweeteners, flavorings, preservatives, and antioxidants may also be present in the pharmaceutical composition. Some examples of pharmaceutically acceptable antioxidants include (1) water-soluble antioxidants such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfite, sodium metabisulfite, and sodium sulfite; (2) oil-soluble antioxidants such as ascorbic acid palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, and alpha-tocopherol; and (3) metal chelating agents such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, and phosphoric acid. In some embodiments, the pharmaceutical formulation includes excipients selected from, for example, cellulose, liposomes, micellar-forming agents (e.g., bile acids), and polymer carriers, such as polyesters and polyanhydrides. The suspension may contain, in addition to the active compound, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxylate, bentonite, agar, and tragacanth, as well as mixtures thereof. Various antibacterial and antifungal agents, such as parabens, chlorobutanol, and phenolsorbic acid, can be included to reliably prevent microbial action of the active compound. It may also be desirable to include isotonic agents such as sugars and sodium chloride in the composition. Furthermore, the inclusion of absorption-delaying agents such as aluminum monostearate and gelatin may provide sustained absorption of the injectable pharmaceutical form.

[0194] In synthetic chemistry, a “protecting group” or “protecting group” means a group that selectively blocks one reactive site in a polyfunctional compound so that a chemical reaction can selectively occur at another unprotected reactive site. Certain processes in this disclosure rely on a protecting group blocking reactive nitrogen and / or oxygen atoms present in the reactants. For example, the terms “amino protecting group” and “nitrogen protecting group” are used synonymously herein, and these terms refer to organic groups intended to protect nitrogen atoms from undesirable reactions during a synthetic procedure. Examples of nitrogen protecting groups include, but are not limited to, trifluoroacetyl, acetamide, benzyl (Bn), benzyloxycarbonyl (carbobenzyloxy, CBZ), p-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl, and tert-butoxycarbonyl (BOC). Those skilled in the art will understand how to select a group in terms of ease of removal and ability to withstand subsequent reactions.

[0195] "Subjects" means mammals and non-mammals. Mammals mean any member of the class Mammalia, such as humans; non-human primates, e.g., chimpanzees and other apes, as well as monkey species; pastoral animals, e.g., cattle, horses, sheep, goats, and pigs; domestic animals, e.g., rabbits, dogs, and cats; laboratory animals, including rodents, e.g., rats, mice, and guinea pigs. Examples of non-mammals include birds, etc. The term "subjects" does not indicate a specific age or sex.

[0196] "Therapeutic dose" refers to the amount of compound sufficient to exert the therapeutic effect on a diseased condition when administered to a subject for that purpose. The therapeutic dose will vary depending on the compound, the diseased condition being treated, the severity of the disease, the subject's age and relative health status, the route and form of administration, the judgment of the attending physician or veterinarian, and other factors.

[0197] The terms “as defined above” and “as defined herein” incorporate, by reference, the broader definition of a variable, as well as any preferred, less preferred, and most preferred definitions, if any, of that variable.

[0198] "To treat" a disease state or "treatment" of a disease state includes (i) preventing the disease state, i.e., preventing the development of the clinical symptoms of a disease state in a person who is likely to develop or be predisposed to the disease state but has not yet experienced or shown symptoms of the disease state; (ii) inhibiting the disease state, i.e., suppressing the development of the disease state or its clinical symptoms; or (iii) alleviating the disease state, i.e., causing a temporary or permanent recession of the disease state or its clinical symptoms.

[0199] As used herein, the phrase “each may be identical or different” (or similar variations thereof) means that each description (i.e., “instance”) of a single-group variable in the general formula (e.g., “L1” or “R3”) may be the same as or different from the other of that variable, for example, each instance may be independently selected from a set or list of options available in that group. Thus, in a particular embodiment of the general formula, two positions labeled with the same variable may both have different selections or values. As a non-limiting example, embodiments of the general structure AX may include embodiments in which three of the R3 groups of CD are hydrogen and four of the R3 groups of CD are 2-hydroxypropyl.

[0200] II. Compounds

[0201] This disclosure describes the design and testing of various dimers of CD. As shown in Figure 1A, CD is a cyclic oligosaccharide composed of D-glucose molecules of the unmodified (i.e., unsubstituted) state of 6(αCD), 7(βCD), or 8(γCD). These CDs can be substituted in a variety of ways, including, but not limited to, functionalization of the hydroxyl groups at the C2, C3, and C6 positions of the glucose ring with methyl, succinyl, sulfobutyl, or hydroxypropyl groups, as shown in Figure 1B, which shows the chemical structure of an exemplary HPβCD having substitutions at various positions around the CD ring.

[0202] As shown in Figure 3A, it is sometimes convenient to describe CD as having a "frustum of a cone" shape with a primary (1°) plane and a secondary (2°) plane. The primary and secondary planes of the CD monomer can be described as the "small" plane and the "large" plane, respectively, due to the difference in the number and orientation of hydroxyl groups available on each plane in the unsubstituted state. The oxygen at position 4 of each D-glucose monomer (i.e., the "O4" oxygen) can be considered to form a ring around the frustum of a cone between the primary and secondary planes.

[0203] As shown in Figure 3B, where the head and tail groups are indicated, the CD monomer engages in host-guest chemistry with biomolecules such as 7-ketocholesterol (7KC) to form a 7KC-CD complex. As shown in Figure 3C, when 7KC forms a complex with CD, the principal axis of 7KC forms an angle with respect to the axis perpendicular to the plane of the O4 atom ring. Furthermore, 7KC can form a complex with CD using either the head or tail group that is involved in the primary plane. As shown in Figure 3D (top), when the tail group is involved in the primary plane, 7KC can be said to be oriented downwards. As shown in Figure 3D (bottom), when the head group is involved in the primary plane, 7KC can be said to be oriented upwards.

[0204] CD dimers can be produced by linking two CD monomers with one or more linking groups. CD dimers composed of CD monomers with different ring sizes may be considered "heterodimers." CD dimers composed of CD monomers with identical ring sizes may be considered "homodimers" or "asymmetric dimers," depending on whether their substituents are the same or different. These dimers may include, but are not limited to, HPα-βCD dimers, methyl-α-βCD dimers, succinyl-α-βCD dimers, sulfobutyl-α-βCD dimers, HPβCD dimers, methyl-βCD dimers, succinyl-βCD dimers, sulfobutyl-βCD dimers, and quaternary ammonium dimers (e.g., 2-hydroxytrimethylammoniumpropyl). Figures 7C, 7H, 7M, 7S, 7AC, and 7AI show various functionalized CD dimers disclosed herein. The inventors previously described how the affinity of certain dimers for 7KC and cholesterol is dramatically increased compared to the monomeric CD. Figure 3A shows an exemplary CD dimer complexed with a sterol in an empty state, in contrast to the CD monomer complexed in an empty state in Figure 3D. This disclosure describes dimers including specific combinations of CD monomers, the exemplary embodiments of which have enhanced binding properties.

[0205] In certain embodiments, two or more chemical formula variables or each pair may be considered "condensed" because each variable may be an "identical" selection of another variable, or the identical variable may be another. By the term "condensed" as applied to chemical formula variables and variable instances, it should be understood that two or more variables and / or each pair are connected such that there is a continuous chain of atoms between any two atoms of the "condensed" variable, without passing through any atom not represented by the "condensed" variable or each pair. As a non-limiting example, a person skilled in the art will understand that divalent substituents connected to two R1s of a CD structure (e.g., structures A-Xa, A-Xb, B-Xa, B-Xb, C-Xa, C-Xb, etc.) may be considered an "identical" selection such that they "condense" with respect to each R1. As another non-limiting example, a person skilled in the art will understand that the structures of CD (e.g., structures A-Xa, A-Xb, B-Xa, B-Xb, C-Xa, C-Xb, etc.) and the divalent substituents connected to one R1 and the other R2 may be considered "identical" choices such that they are "condensed" with respect to one R1 and the other R2.

[0206] The set of available choices for the variables mentioned above as part of a particular embodiment of General Formula I means that a particular choice from an adjacent variable may result in redundant choices and embodiments. For example, a person skilled in the art will understand that the choice of bond and hydroxyl group for each L1 / R1 pair is identical to the choice of -O- and hydrogen for each pair. A person skilled in the art will also understand that it is not necessary to enumerate all possible redundant choices across each variable in order to present any particular embodiment. Furthermore, if it is stated that a particular structure does not exist for a particular combination of variable values, it means that other combinations of variable values ​​that produce that structure are also prohibited. For example, if it is stated that the L1 / R1 pair may not be oxygen and hydrogen, it also means that L1 and R1 may not be bond and hydroxyl, for assigning these values ​​results in the exact same structure that is identified as not existing.

[0207] The compounds disclosed herein are intended to include any combination of isotopic isomers that allow for the selection of groups herein. Furthermore, where stereochemistry exists or where one or more stereoisomers can be generated by the selection of various groups, it should be understood that the disclosure herein includes both racemic mixtures and isolated stereoisomer products unless otherwise specified. In preferred embodiments, each CD monomer contains all D-glucose.

[0208] In one embodiment, the present disclosure provides a CD dimer of the general structure CD-L-CD' in which one or both of CD and CD' are specifically and completely substituted at the C6 position (i.e., having a choice for L3 / R3 and L3' / R3' which are not bonded and hydroxyl, respectively). Such a position may also be said to be “saturated”. Although not intended to be limited by theory, it is thought that by placing the substitution only at the C6 position, the hydrophobic cavities of one or both of CD and CD' can be effectively expanded, thereby creating a better environment for encapsulation of the 7KC tail group with a long aliphatic chain and other sterols. By substituting only on the primary face of CD, all unmodified hydroxyl groups on the secondary face are utilized for hydrogen bonding with the head group of the target molecule and the hydroxyl group of CD on the opposite side, and the stability of the complex is improved by both mechanisms. An additional potential advantage of C6 substitution is that, in some cases, they can be made as single isomer molecules. This may reduce complexity and improve batch-to-batch reproducibility of the final product.

[0209] In another embodiment, the disclosure provides CD dimers in which alkyl groups are used as substituents. Although not intended to be limited by theory, alkyl groups are considered to have a greater ability to expand one or both hydrophobic cavities of the subunit because they are more hydrophobic than charged and polar substitutions, and thus expand, creating a better environment for encapsulation of other sterols having a 7KC tail group and a long aliphatic chain.

[0210] This disclosure includes further substitution of the dimerized CD described herein (e.g., HPβCD or another CD). Chemical modification may be performed before or after dimerization. Chemical modification of CD can be performed directly on the unmodified beta-CD ring or on CD with appropriately introduced functional groups by reacting with a chemical reagent (nucleophile or electrophile) (Adair-Kirk [et al.], Nat. Med., 14(10):1024-5, (2008)); (Khan, [et al.], Chem. Rev., 98(5):1977-1996, (1998)). To date, more than 1,500 CD derivatives have been prepared by chemical modification of unmodified CD. CD can also be prepared by de novo synthesis starting from an oligopyranoside linked to glucopyranose. Such synthesis can be achieved using various chemical reagents or biological enzymes, such as CD transglycosylase. An overview of chemically modified CDs as drug carriers in drug delivery systems is provided, for example, in (Stella, [et al.], Toxicol. Pathol., 36(1):30-42, (2008)), the disclosure of which is incorporated herein by reference in its entirety. U.S. Patents 3,453,259 and 3,459,731 describe electrically neutral CDs, the disclosures of which are incorporated herein by reference in their entirety. Other derivatives include cationic CDs disclosed in U.S. Patent 3,453,257; insoluble crosslinked CDs disclosed in U.S. Patent 3,420,788; and anionic CDs disclosed in U.S. Patent 3,426,011, all of which are incorporated herein by reference in their entirety. Among the anionic CD derivatives, carboxylic acids, phosphorous acids, phosphinoic acids, phosphonic acids, phosphoric acids, thiophosphonic acids, thiosulfinic acids, and sulfonic acids have been added to the parent CD, as disclosed, for example, in U.S. Patent No. 3,426,011. Sulfoalkyl ether CD derivatives are also described, for example, in U.S. Patent No. 5,134,127, which is incorporated herein by reference in its entirety.In some embodiments, cyclic oligosaccharides can have two or more monosaccharide units substituted with a triazole ring, which can be synthesized by azido-alkyne hysgen cycloaddition (Bodine [et al.], J. Am. Chem. Soc., 126(6):1638-9, (2004)).

[0211] The two CD monomers of the CD dimer of this disclosure are linked by a linker (also referred to herein as a linking group). Methods available for linking the CD subunit to the linker are described below. Further methods for linking the CD subunit to the linker are known in the art (Georgeta [et al.], J. Bioact. Compat. Pol., 16:39-48. (2001)), (Liu [et al.], Acc. Chem. Res., 39:681-691. (2006)), (Ozmen [et al.], J. Mol. Catal. B-Enzym., 57:109-114. (2009)), (Trotta [et al.], Compos. Interface, 16:39-48. (2009)), each of which is incorporated herein by reference in whole. For example, a linker group containing a moiety that reacts with a hydroxyl group (e.g., a carboxyl group that can be activated by carbodiimide) can be reacted with CD to form a covalent bond. In another example, one or more hydroxyl groups of CD can be activated by a known method (e.g., tosylation) and reacted with a reactive group of the linker (e.g., an amino group).

[0212] Generally, a linker has two reaction moieties that first react with and bond to each CD monomer. In one embodiment, the linker is first bonded to the CD to produce a linker-CD compound, which is then isolated. The remaining reaction moiety of the linker in the linker-CD compound is then subsequently reacted with a second CD. The linker-cyclodextrin compound may be further modified with protecting groups and / or ad hoc-designed functional groups to introduce additional interacting functional groups and / or to achieve the desired regiochemistry at the target key intermediate. The second reaction moiety of the linker may be protected while the first reaction group reacts, but may not be protected if the first reaction moiety of the linker and the first reaction moiety react with the two molecules differently. The linker may react with both molecules simultaneously and link them together. In another embodiment, the linker may have additional reaction groups for linking with other molecules.

[0213] Several linkers are known in the art. Such linkers can be used to link any of the various groups together, when the various groups possess or have been introduced with functional groups that can react with the reactive linker to link. Some groups that can react with a double reactive linker include amino, thiol, hydroxyl, carboxyl, ester, and alkyl halide groups. For example, cyclic oligosaccharides and polysaccharides can be linked using amino-amino coupling reagents when each group to be linked possesses at least one amino group. Some examples of amino-amino coupling reagents include diisocyanate, alkyl dihalides, dialdehydes, disuccinimidyl suberate (DSS), disuccinimidyl tartrate (DST), and disulfosuccinimidyl tartrate (sulfo-DST), all of which are commercially available. In another embodiment, an amino-thiol coupling agent can be used to link a thiol group of one molecule to an amino group of another molecule. Some examples of amino-thiol coupling reagents include succinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-carboxylic acid (SMCC) and sulfosuccinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-carboxylic acid (sulfo-SMCC). In yet another embodiment, a thiol-thiol coupling agent can be used to link groups having at least one thiol group.

[0214] In some embodiments, the linker is so small that its length is only one atom (e.g., --O--, --CH2--, or --NH-- linkage), or two or three atoms (e.g., amide, ureid, carbamate, ester, carbonate, sulfone, ethylene, or trimethylene linkage). In other embodiments, the linker provides a higher degree of freedom of motion by having an atomic length of at least 4, 5, 6, 7, or 8 atoms, up to a maximum of, for example, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, or 30 atoms. Preferred linker lengths are 2 to 12 atoms, or 4 to 8 atoms. In exemplary embodiments, the linker is a C4 alkyl, which may be unsubstituted. In some embodiments, the linker comprises a triazole (e.g., B is a triazole). In further embodiments, the linker comprises a triazole linked to each CD monomer by alkyl chains of equal or different lengths (for example, A and A' are alkyl chains of varying lengths, and B is the triazole).

[0215] In another embodiment, the disclosure provides a method for producing CD dimers that have specificity for other smaller hydrophobic molecules. An exemplary method is carried out by first producing a CD dimer nucleus having a specific structure to which specificity is conferred by synthesis. Then, while maintaining the high affinity conveyed by the CD dimer nucleus, any substitutions can be made to create specificity for selected hydrophobic molecules. This specificity can be further modified with different linkers.

[0216] Mai. Pharmaceutical composition

[0217] In another embodiment, the Disclosure provides a pharmaceutical composition comprising a CD dimer composition and a pharmaceutically acceptable carrier disclosed herein. The pharmaceutical composition may be suitable for administration to a target, for example, parenterally (e.g., subcutaneous, intramuscular, or intravenous), topically, transdermally, or orally, sublingually, or buccally, preferably intravenously or subcutaneously, more preferably intravenously. The CD dimer composition may be the sole active ingredient in the composition. The pharmaceutical composition may consist of, or essentially consist of, the CD dimer and the pharmaceutically acceptable carrier.

[0218] In another embodiment, the Disclosure provides a pharmaceutical composition comprising one or more CD dimers and a hydrophobic drug disclosed herein. The hydrophobic drug may include hormones or sterols, such as estrogen, estrogen analogs, etc. The one or more CD dimers may be present in an amount effective to solubilize the hydrophobic drug.

[0219] The term "pharmaceutically acceptable" is used herein to refer to a compound, material, composition, and / or dosage form that, within the bounds of sound medical judgment, is suitable for entry into a living organism or living biological tissue, and preferably, is free from significant toxicity, irritation, or allergic reactions. The present invention comprises a method comprising administering a CD dimer to a patient, wherein the CD dimer is contained within a pharmaceutical composition. The pharmaceutical compositions of the present invention are formulated with pharmaceutically acceptable carriers, excipients, and other agents that provide suitable transport, delivery, tolerance, etc. Numerous suitable formulations can be found in formularies known to pharmaceutical chemists, e.g., Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa. Examples of these formulations include powders, pastes, ointments, gels, waxes, oils, lipids, lipid (cationic or anionic)-containing vesicles (e.g., LIPOFECTIN®), DNA complexes, anhydrous absorbent pastes, oil-in-water and water-in-oil emulsions, carbowax emulsions (polyethylene glycol of various molecular weights), semi-solid gels, and carbowax-containing semi-solid mixtures. (See also Powell [et al.], J. Pharm. Sci. Technol., 52:238-311, (1998)).

[0220] When used herein, the term “pharmaceutically acceptable carrier” generally refers to a pharmaceutically acceptable composition useful for introducing an active agent into the body, such as a liquid or solid filler, diluent, excipient, manufacturing aid (e.g., lubricant, magnesium talc, calcium stearate or zinc stearate, or stearic acid), or solvent encapsulation material. Each carrier must be “acceptable” in the sense that it is compatible with the other components of the formulation and is not harmful to the patient. Examples of suitable aqueous and non-aqueous carriers usable in the pharmaceutical compositions of the present invention include, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.), vegetable oils (e.g., olive oil), and organic esters for injection (e.g., ethyl oleate), as well as suitable mixtures thereof. Adequate fluidity can be maintained, for example, by the use of a coating material such as lecithin, by maintaining the required particle size in the case of dispersion, and by the use of a surfactant.

[0221] Other examples of materials that can function as pharmaceutically acceptable carriers include: (1) sugars, e.g., lactose, glucose, and sucrose; (2) starches, e.g., corn starch and potato starch; (3) cellulose and its derivatives, e.g., sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, e.g., cocoa butter and suppository wax; (9) oils, e.g., peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; (10) glycols. Examples include (11) propylene glycol, polyols such as glycerin, sorbitol, mannitol, and polyethylene glycol, (12) esters such as ethyl oleate and ethyl laurate, (13) agar, (14) buffers such as magnesium hydroxide and aluminum hydroxide, (15) alginic acid, (16) pyrogen-free water, (17) isotonic saline, (18) Ringer's solution, (19) ethyl alcohol, (20) pH buffer, (21) polyesters, polycarbonates, and / or polyanhydrides, and (22) other non-toxic and suitable substances used in pharmaceutical formulations.

[0222] Various auxiliary agents such as wetting agents, emulsifiers, lubricants (e.g., sodium lauryl sulfate and magnesium stearate), colorants, release agents, coating agents, sweeteners, flavorings, preservatives, and antioxidants may also be present in the pharmaceutical composition. Some examples of pharmaceutically acceptable antioxidants include (1) water-soluble antioxidants such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfite, sodium metabisulfite, and sodium sulfite; (2) oil-soluble antioxidants such as ascorbic acid palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, and alpha-tocopherol; and (3) metal chelating agents such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, and phosphoric acid. In some embodiments, the pharmaceutical formulation includes excipients selected from, for example, cellulose, liposomes, micellar-forming agents (e.g., bile acids), and polymer carriers, such as polyesters and polyanhydrides. The suspension may contain, in addition to the active compound, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxylate, bentonite, agar, and tragacanth, as well as mixtures thereof. Various antibacterial and antifungal agents, such as parabens, chlorobutanol, and phenolsorbic acid, can be included to reliably prevent microbial action of the active compound. It may also be desirable to include isotonic agents such as sugars and sodium chloride in the composition. Furthermore, the inclusion of absorption-delaying agents such as aluminum monostearate and gelatin may provide sustained absorption of the injectable pharmaceutical form.

[0223] The pharmaceutical formulations of the present invention can be prepared by any method known in the pharmaceutical art. The amount of the active ingredient (i.e., a CD dimer such as HPβCD dimer or another CD dimer of this disclosure) that can be combined with a carrier material to make a single dosage form varies depending on the host being treated and the specific mode of administration. The amount of the active ingredient that can be combined with a carrier material to make a single dosage form is generally the amount of compound that produces the therapeutic effect. The amount of the active compound may range from about 0.1 to 99.9 percent, more typically from about 80 to 99.9 percent, and more typically from about 99 percent. The amount of the active compound may range from about 0.1 to 99 percent, more typically from about 5 to 70 percent, and more typically from about 10 to 30 percent. In exemplary embodiments, dosage forms for intravenous administration are provided, which are aqueous solutions with a concentration of 0.5% to 0.001%, for example, 0.12% to 0.0105%, for example, about 0.01% (W / V). In exemplary embodiments, dosage forms for intravenous administration are provided, which are aqueous solutions with concentrations of 2.5% to 0.25%, for example, 2% to 0.5%, for example, about 1% (W / V). In exemplary embodiments, the dosage form provides intravenous administration of up to 500 mL of a 1% (W / V) solution, which results in a maximum dose of 5 grams.

[0224] Formulations of the present invention suitable for oral administration may be in the form of capsules, cachets, pills, tablets, sweetened tablets (using flavoring agents, usually sucrose and acacia or tragacanth), powders, granules, or solutions or suspensions in aqueous or non-aqueous liquids, or oil-in-water or water-in-oil liquid emulsions, or elixirs or syrups, or pastils (using inert agents such as gelatin and glycerin, or sucrose and acacia), and / or mouthwashes, each containing a predetermined amount of the compound of the present invention as the active ingredient. The active compound may also be administered as a bolus, lick, or paste.

[0225] The preparation methods for these formulations or compositions generally involve the step of mixing the compound of the present invention with a carrier and optionally one or more auxiliary agents. In the case of solid dosage forms (e.g., capsules, tablets, pills, powders, granules, lozenges, etc.), the active compound can be mixed with a pulverized solid carrier and molded, typically by pelletizing, tableting, granulating, powdering, or coating. Generally, examples of solid carriers include sodium citrate or dicalcium phosphate and / or any of the following: (1) Fillers or bulking agents, e.g., starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; (2) Binders, e.g., carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and / or acacia; (3) Humectants, e.g., glycerol; (4) Disintegrants, e.g., agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; (5) Dissolution retarders, e.g., paraffin; (6) Absorption enhancers, e.g., quaternary ammonium compounds and ammonium compounds. (7) surfactants, such as poloxamer and sodium lauryl sulfate; (8) wetting agents, such as cetyl alcohol, glycerol monostearate, and nonionic surfactants; (9) absorbents, such as kaolin and bentonite clay; (10) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, zinc stearate, sodium stearate, stearic acid, and mixtures thereof; (11) colorants; and (22) release control agents, such as crospovidone or ethylcellulose. In the case of capsules, tablets, and pills, the pharmaceutical composition may also contain buffers. Similar types of solid compositions may also be used as fillers in soft and hard shell gelatin capsules, using excipients such as lactose and high molecular weight polyethylene glycol.

[0226] Tablets may be manufactured by compression or molding with one or more auxiliary components as optional. Compressed tablets may be prepared using binders (e.g., gelatin or hydroxypropyl methylcellulose), lubricants, inert diluents, preservatives, disintegrants (e.g., sodium starch glycolate or cross-linked sodium carboxymethylcellulose), surfactants, or dispersants.

[0227] Tablets and other solid dosage forms of activators, such as capsules, pills, and granules, can optionally be surface-treated or prepared using coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical manufacturing field. These dosage forms can also be formulated to release the active ingredient in a sustained or controlled manner, for example, by using hydroxypropyl methylcellulose in varying proportions to achieve a desired release profile, along with other polymer matrices, liposomes, and / or microspheres. Alternatively, the dosage forms may be formulated for rapid release, for example, by lyophilization.

[0228] In general, dosage forms need to be sterilized. For this purpose, dosage forms can be sterilized, for example, by filtration with a bacterial collection filter, or by incorporating a sterilizing agent in the form of a sterile solid composition that can be dissolved in sterile water or some other sterile injection medium immediately before use. Pharmaceutical compositions may contain opacifiers and may release the active ingredient(s) in a manner that is selectively delayed only in or preferentially in specific parts of the gastrointestinal tract. Examples of embedding compositions that can be used include polymers and waxes. The active ingredient(s) may also be in a microencapsulated form together with one or more of the excipients described above, where appropriate.

[0229] Liquid dosage forms are typically pharmaceutically acceptable emulsions, microemulsifies, solutions, suspensions, syrups, or elixirs of the active ingredient. In addition to the active ingredient, liquid dosage forms may include, for example, water or other solvents, solubilizers and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerol, tetrahydrofuryl alcohol, polyethylene glycol and sorbitan fatty acid esters, and mixtures thereof, which are commonly used in the art.

[0230] Dosage forms specifically intended for topical or transdermal administration may be, for example, in the form of powders, sprays, ointments, pastes, creams, lotions, gels, solutions, or patches. Ophthalmic formulations, such as ophthalmic ointments, powders, solutions, etc., are also intended herein. The active compound may be mixed under sterile conditions with a pharmaceutically acceptable carrier and any preservatives, buffers, or propellants as needed. In addition to the active compound of the present invention, topical or transdermal dosage forms may contain one or more excipients, such as animal and vegetable fats, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicones, bentonite, silicic acid, talc, and zinc oxide, and mixtures thereof. Sprays may also contain common propellants, such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, such as butane and propane.

[0231] With regard to the object of the present invention, transdermal patches may offer the advantage of being able to deliver the compounds of the present invention to the body in a controlled manner. Such dosage forms can be prepared by dissolving or dispersing the compounds in a suitable culture medium. Absorption enhancers may also be included to increase the flow of the compounds across the skin. Such flow rates can be controlled by either providing a rate-controlled membrane or dispersing the compounds in a polymer matrix or gel.

[0232] A pharmaceutical composition of the present invention suitable for parenteral administration generally comprises one or more compounds of the present invention in combination with one or more pharmaceutically acceptable sterile isotonic aqueous or nonaqueous solutions, dispersions, suspensions, or emulsions, or a sterile powder that can be reconstituted into a sterile injection solution or dispersion before use, the sterile powder of which may contain sugars, alcohols, antioxidants, buffers, bacteriostatic agents, or solutes that make the formulation isotonic with the blood of the target recipient.

[0233] In some cases, it may be desirable to delay drug absorption from subcutaneous or intramuscular injection in order to prolong the drug's effect. This can be achieved by using liquid suspensions of crystalline or amorphous materials with poor water solubility. The rate of drug absorption may depend on the rate of dissolution and, therefore, on the crystal size and crystal form. Alternatively, the delay of absorption of parenterally administered dosage forms can be achieved by dissolving or suspending the drug in an oily vehicle.

[0234] Depot injection formulations can be prepared by forming a microencapsulation matrix in which the active compound is contained in a biodegradable polymer such as polylactide-polyglycolide. The drug release rate can be controlled depending on the ratio of the drug to the polymer and the properties of the specific polymer used. Examples of other biodegradable polymers include poly(orthoester) and poly(anhydride). Depot injection formulations can also be prepared by capturing the drug in liposomes or microemulsifies that are compatible with body tissues.

[0235] Pharmaceutical compositions may be in the form of microemulsions. In the form of microemulsions, the bioavailability of the active ingredient may be improved. See (Dorunoo [et al.], Drug Development and Industrial Pharmacy, 17(12):1685-1713 (1991)) and (Sheen [et al.], J. Pharm. Sci., 80(7):712-714, (1991)). The contents of these documents are incorporated herein by reference in their entirety.

[0236] The pharmaceutical composition may contain micelles formed from the compound of the present invention and at least one amphiphilic carrier, in which case the micelles have an average diameter of less than about 100 nm. In some embodiments, the micelles have an average diameter of less than about 50 nm, or an average diameter of less than about 30 nm, or an average diameter of less than about 20 nm.

[0237] Although any suitable amphiphilic carrier is considered herein, amphiphilic carriers are generally those that have been approved as inactive pharmaceutical components and are capable of both dissolving the compounds of the present invention and microemulsifying them when the solution comes into contact with a complex aqueous phase (e.g., those found in living biological tissues) in a subsequent step. Typically, amphiphilic components that meet these requirements have an HLB (hydrophilic-to-lipophilic balance) value of 2 to 20 and their structure contains linear aliphatic radicals in the C-6 to C-20 range. Some examples of amphiphilic agents include polyethylene glycolated aliphatic glycerides and polyethylene glycols.

[0238] Particularly preferred amphiphilic carriers are saturated and monounsaturated polyethylene glycolated fatty acid glycers, for example, those obtained by fully or partially hydrogenating various vegetable oils. These oils may advantageously consist of tri, di, and mono fatty acid glycers, as well as di and monopolyethylene glycol esters of the corresponding fatty acids. Particularly preferred fatty acid compositions include 4%-10% capric acid, 3%-9% capric acid, 40%-50% lauric acid, 14%-24% myristic acid, 4%-14% palmitic acid, and 5%-15% stearic acid. Another useful class of amphiphilic carriers includes partially esterified sorbitan and / or sorbitol using saturated or monounsaturated fatty acids (SPAN series) or their corresponding ethoxylated analogs (TWEEN series). Commercially available amphiphilic carriers are particularly intended, and examples of such carriers include the Gelucire® series, Labrafil®, Labrasol®, or Lauroglycol®, PEG-monooleate, PEG-dioleate, PEG-monolaurate and dilaurate, lecithin, and polysorbate 80.

[0239] IV. Indications for the disease, and methods of treatment and prevention.

[0240] An exemplary embodiment of the present invention provides the use of a CD dimer as disclosed herein for solubilizing and / or removing 7KC, which can be done both in vitro and in vivo.

[0241] In exemplary embodiments, the CD dimer disclosed herein exhibits higher binding affinity and / or solubilization to 7KC than to cholesterol. While the specificity of 7KC to cholesterol is most pronounced at near-saturated concentrations, solubilization of both sterols at higher concentrations may approach 100%. This specificity allows for the use of such CD dimers to preferentially solubilize and remove 7KC.

[0242] 7KC is thought to be involved in heart disease, cystic fibrosis, liver damage and failure, and complications of hypercholesterolemia. In individuals with hypercholesterolemia, 7KC may diffuse across cell membranes, in which case it affects receptor and enzyme function. The high rate of dementia in hypercholesterolemia has been linked to 7KC accumulation. In the liver, 7KC affects tissue fenestration and porosity, which increase with age. 7KC also promotes the migration of cytoplasmic NADPH oxidase components to neutrophil (leukocyte) membranes and enhances rapid reactive oxygen species production. The pathogenesis of other age-related diseases, such as age-related macular degeneration (AMD-atrophic type), Alzheimer's disease, and lysosomal storage disorders like Niemann-Pick disease type C (NPC), has also been linked to elevated 7KC levels. Oxysterols, including 7KC, are also involved in elevated free radical levels, which in turn affect lipid circulation in cystic fibrosis. Increased free radicals induced by oxysterols such as 7KC are thought to be involved in apoptosis, cytotoxicity, impaired endothelial function, and regulation of enzymes involved in inflammation and fatty acid metabolism.

[0243] 7KC is formed from a non-enzymatic reaction between oxygen radicals and cholesterol, which suggests that 7KC formation may not be beneficial. In fact, 7KC is thought to enhance free radical production throughout the body, but this is of particular concern in cardiovascular tissue. Free radicals affect cellular and enzymatic reactions that are important for cholesterol-mediated tissue damage, and this is especially critical in these tissues. Free radicals are thought to enhance inflammation in vascular structures. 7KC is thought to cause mitochondrial and lysosome dysfunction by disrupting the function of cell and organelle membranes, and is thought to be involved in increasing the frequency of foam cell formation from macrophages in atherosclerotic plaques. While the scavenging function of these macrophages is expected to help alleviate plaque, if they are filled with cholesterol and oxysterols, they may not, and instead become part of the plaque.

[0244] Exemplary embodiments provide treatment for diseases associated with and / or exacerbated by the accumulation of 7KC, including atherosclerosis, AMD, arteriosclerosis, coronary atherosclerosis due to coronary artery calcification lesions, heart failure (all stages), Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, Huntington's disease, vascular dementia, multiple sclerosis, Smith-Lemle-Oppitz syndrome, infantile neuronal ceroid lipofuscinosis, and lysosomal acid lipase. Examples include deficiencies, cerebral tendon xanthomatous dystrophy, X-linked adrenoleukodystrophy, sickle cell disease, Niemann-Pick disease type A, Niemann-Pick disease type B, Niemann-Pick disease type C, Gaucher disease, Stargardt disease, idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, cystic fibrosis, liver injury, liver failure, non-alcoholic steatohepatitis, non-alcoholic fatty liver disease, irritable bowel syndrome, Crohn's disease, ulcerative colitis, and / or hypercholesterolemia or dementia associated with hypercholesterolemia. Preferred CD dimers are selective for 7KC (more so than cholesterol). Preferably, the above CD dimers preferentially solubilize 7KC while minimizing or avoiding potentially harmful or toxic effects that may result from excessive cholesterol removal.

[0245] In another embodiment, the Disclosure provides a therapy comprising administering an effective amount of the CD dimer composition as disclosed herein to a subject in need thereof, the subject may be suffering from adverse or toxic effects of 7KC or symptoms associated with adverse or toxic effects of 7KC.

[0246] In another embodiment, the Disclosure provides a method for reducing the amount of 7KC in a subject requiring a reduction in the amount of 7KC, comprising administering to the subject in an effective amount of a CD dimer composition as disclosed herein or a pharmaceutical composition comprising a CD dimer composition as disclosed herein.

[0247] The CD dimer composition can be administered to the subject parenterally (e.g., subcutaneously, intramuscularly, or intravenously), topically, transdermally, or orally, sublingually, or buccally, preferably intravenously.

[0248] The method may include (a) administering the CD dimer composition to the subject in an amount of about 1 mg to 20 g, for example, 10 mg to 1 g, 50 mg to 200 mg, or 100 mg; (b) administering the CD dimer composition to the subject in an amount of about 1 to 10 g, for example, about 2 g, about 3 g, about 4 g, or about 5 g; or (c) administering the CD dimer composition to the subject in an amount of 50 mg to 5 g, for example, 100 mg to 2.5 g, 100 mg to 2 g, or 250 mg to 2.5 g.

[0249] This method is used for atherosclerosis / coronary artery disease, arteriosclerosis, coronary artery atherosclerosis due to coronary artery calcification, heart failure (all stages), Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, Huntington's disease, vascular dementia, multiple sclerosis, Smith-Lemle-Oppitz syndrome, pediatric neuronal ceroid lipofuscinosis, lysosomal acid lipase deficiency, cerebral tendon xanthomatous dystrophy, X-linked adrenoleukodystrophy, sickle cell anemia, Niemann-Pick disease type A, and Niemann-Pick disease. It may be used to prevent, treat, or induce remission of one or more of the following conditions: type B hepatitis, Niemann-Pick disease type C, Gaucher disease, Stargardt disease, age-related macular degeneration (atrophic type), idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, cystic fibrosis, liver injury, liver failure, non-alcoholic steatohepatitis, non-alcoholic fatty liver disease, irritable bowel syndrome, Crohn's disease, ulcerative colitis, and / or hypercholesterolemia. The treatment may be administered selectively in combination with other treatments. The method may include administering secondary treatments to the subject, which may be administered simultaneously or sequentially.

[0250] The method may be used to prevent, treat, or alleviate the symptoms of atherosclerosis. The CD dimer composition may be administered in combination with other treatments for the treatment or prevention of atherosclerosis, such as anticholesterol agents, antihypertensive agents, antiplatelet agents, nutritional supplements, or surgery or behavioral interventions, including, but not limited to, those described herein. The anticholesterol agents may include fibrates or statins, antiplatelet agents, antihypertensive agents, or nutritional supplements. The statin may include ADVICOR(R) (sustained-release niacin / lovastatin), ALTOPREV(R) (sustained-release lovastatin), CADUET(R) (amlodipine / atorvastatin combination), CRESTOR(R) (rosuvastatin), JUVISYNC(R) (sitagliptin / simvastatin), LESCOL(R) (fluvastatin), LESCOL XL (sustained-release fluvastatin), LIPITOR(R) (atorvastatin), LIVALO(R) (pitavastatin), MEVACOR(R) (lovastatin), PRAVACHOL(R) (pravastatin), SIMCOR(R) (sustained-release niacin / simvastatin), VYTORIN(R) (ezetimibe / simvastatin), or ZOCOR(R) (simvastatin).

[0251] The method may be used to prevent, treat, or induce remission of symptoms of atrophic age-related macular degeneration. The method may be used to prevent, treat, or induce remission of symptoms of Stargardt disease. The CD dimer composition may be administered in combination with other treatments for the treatment or prevention of atrophic AMD or Stargardt disease, such as the AREDS nutritional supplement formulation containing LBS-008 (Belite Bio) (a non-retinoid antagonist of retinol-binding protein 4), vitamin C and vitamin E, beta-carotene, zinc, and copper; the AREDS2 nutritional supplement formulation containing vitamin C and vitamin E, zinc, copper, lutein, zeaxanthin, and omega-3 fatty acids; or a combination thereof.

[0252] The method may be used to prevent, treat, or alleviate the symptoms of Niemann-Pick disease. The CD dimer composition may be administered in combination with one or more other treatments for the treatment or prevention of Niemann-Pick disease, such as miglustat (ZAVESCA(R)), HPβCD (TRAPPSOL CYCLO, VTS-270), and physiotherapy.

[0253] The method may be used to prevent, treat, or induce remission of symptoms of Alzheimer's disease. The CD dimer composition may be administered in combination with other treatments for the treatment or prevention of Alzheimer's disease, such as cholinesterase inhibitors (ARICEPT(R), EXELON(R), RAZADYNE(R)) and memantine (NAMENDA(R)) or combinations thereof.

[0254] The method may be used to prevent, treat, or alleviate symptoms of heart failure. The CD dimer composition may be administered in combination with other therapies for the treatment or prevention of heart failure, such as one or more aldosterone antagonists, ACE inhibitors, ARBs (angiotensin II receptor blockers), ARNIs (angiotensin receptor neprilysin inhibitors), beta-blockers, vasodilators, calcium channel blockers, digoxin, diuretics, cardiac pump drugs, potassium, magnesium, selective sinoatrial node inhibitors, or combinations thereof.

[0255] In exemplary embodiments, a patient may be administered 1 mg to 10 g of CD dimers, for example, 10 mg to 1 g or 100 mg to 500 mg. In exemplary embodiments, a patient may be administered about 400 mg of CD dimers. In exemplary embodiments, a patient may be administered 1 to 10 g of CD dimers, for example, about 2 g, about 3 g, about 4 g, or about 5 g. In exemplary embodiments, a patient may be administered 50 mg to 5 g of CD dimers, for example, 100 mg to 2.5 g, 100 mg to 2 g, 250 mg to 2.5 g, for example, about 1 g.

[0256] In exemplary embodiments, a single dosage form is provided which may contain the aforementioned amounts of CD dimers, may be packaged for individual administration, and may optionally further contain a pharmaceutically acceptable carrier or excipient. The total amount of CD dimers in the single dosage form may be as presented above, for example, 1 mg to 10 g, e.g., 10 mg to 1 g, 100 mg to 500 mg, 1 to 10 g of CD dimers, 50 mg to 5 g, 100 mg to 2.5 g, 100 mg to 2 g, 250 mg to 2.5 g, e.g., about 1 g, 2 g, about 3 g, about 4 g, or about 5 g.

[0257] CD (e.g., HPβCD as disclosed herein or another CD) dimers may be administered by any suitable means. Preferred routes of administration include parenteral (e.g., subcutaneous, intramuscular, or intravenous), topical, transdermal, oral, sublingual, or buccal. Such administration may also be performed ocularly (e.g., in the form of eye drops), intravitreously, posterior orbitally, subretinally, or subsclerally, which may be preferred in cases of ocular diseases such as AMD.

[0258] The CD (e.g., HPβCD as disclosed herein or another CD) dimer may be administered to a subject or used in vitro, for example, by applying it to cells or tissues isolated from an animal. The cells or tissues can then be introduced into a subject, whether from the subject from which the cells or tissues were isolated or from another individual, the other individual preferably of the same species.

[0259] The subject receiving treatment (i.e., the patient) is typically an animal, generally a mammal, preferably a human. The subject may also be a non-human animal, such as all vertebrates, e.g., mammals, as well as non-human primates, sheep, dogs, cats, cattle, horses, chickens, amphibians, and reptiles. In some embodiments, the subject is livestock, e.g., cattle, pigs, sheep, poultry, and horses, or companion animals, e.g., dogs and cats. The subject may be genetically male or female. The subjects may be of any age, for example, the elderly (generally at least 60, 70, or 80 years old, or older), subjects in the transitional age from adulthood to old age, adults, subjects in the transitional age from pre-adulthood to adulthood, and pre-adults including young people (e.g., from 13 to 16, 17, 18, or 19 years old), children (generally under 13 years old or pre-adolescent), and infants. Furthermore, the subjects can be of any racial group or genotype. Some examples of human racial groups include the Caucasian, Asian, Latin American, African, African American, Native American, Semitic, and Pacific Islander peoples. The method of the present invention may be more suitable for certain racial groups, for example, the Caucasian, particularly Northern European, and Asian racial groups.

[0260] Atherosclerosis

[0261] The exemplary CD dimers described herein are useful for the prevention or treatment of diseases such as atherosclerosis. A combination of a CD dimer and one or more activators, such as those described herein (e.g., antihyperlipidemic agents such as statins), is useful for treating all forms of atherosclerosis, as well as the signs, symptoms, or complications associated with it. Atherosclerosis (also known as atherosclerotic vascular disease or ASVD, and coronary artery disease or CAD) is a condition in which the arterial walls thicken as a result of the accumulation of fatty substances such as cholesterol. Atherosclerosis is a chronic disease that can remain asymptomatic for decades. Atherosclerosis is a syndrome that affects arterial blood vessels, and the chronic inflammatory response in the arterial wall is largely due to the accumulation of macrophage leukocytes. This chronic inflammatory response is thought to be promoted by low-density lipoproteins (plasma proteins that transport cholesterol and triglycerides) if lipids and cholesterol are not adequately removed from macrophages by functional high-density lipoproteins (HDL). Atherosclerosis is generally referred to as hardening of the arteries or accumulation of material in the arteries. This is caused by the formation of multiple plaques within the arteries.

[0262] The pathobiology of atherosclerotic lesions is complex, but generally, stable atherosclerotic plaques tend to be asymptomatic and are abundant in the extracellular matrix and smooth muscle cells, while unstable plaques are abundant in macrophages and foam cells. The extracellular matrix (also known as the fibrous capsule) separating the lesion from the ductus arteriosus lumen is usually fragile and prone to rupture. Rupture of the fibrous capsule exposes thrombus-forming materials such as collagen to the bloodstream, ultimately inducing thrombus formation in the lumen. During formation, intraluminal thrombi can completely occlude the artery (e.g., coronary artery occlusion), but more frequently, they can detach and enter the bloodstream, ultimately occluding smaller downstream branches and causing thromboembolism (e.g., carotid artery thrombus formation often leads to stroke). Apart from thromboembolism, chronic enlargement of atherosclerotic lesions can lead to complete occlusion of the lumen. Chronic progression of lesions is often asymptomatic until the luminal narrowing becomes so severe that it leads to insufficient blood supply to downstream tissues, resulting in ischemia.

[0263] These complications of progressive atherosclerosis are chronic, slowly progressing, and cumulative. In some cases, a soft plaque may suddenly rupture, causing the formation of a blood clot, which rapidly slows or stops blood flow and leads to the death of the tissue supplied by the artery (infarction). Coronary thrombosis of the coronary arteries is also a common complication that can lead to myocardial infarction. Blockage of arteries leading to the brain can lead to stroke. In progressive atherosclerotic disease, claudication can occur due to insufficient blood supply to the legs, which can typically be caused by a combination of stenosis and aneurysmal segments narrowed by blood clots.

[0264] Atherosclerosis can affect all arterial trees, but typically, the risk is higher for larger, high-pressure vessels such as the coronary arteries, renal arteries, femoral arteries, cerebral arteries, and carotid arteries.

[0265] Signs, symptoms, and complications of atherosclerosis include, but are not limited to, elevated plasma total cholesterol, VLDL-C, LDL-C, free cholesterol, cholesterol esters, triglycerides, phospholipids, and arterial lesions (e.g., plaques), as discussed above. In some cases, elevated cholesterol (e.g., total cholesterol, free cholesterol, and cholesterol esters) may be found in one or more of the plasma, aortic tissue, and aortic plaques.

[0266] Certain individuals may have a predisposition to atherosclerosis. Therefore, this disclosure relates to a method of administering a CD dimer alone or in combination with one or more additional therapeutic agents (e.g., antihyperlipidemic agents such as statins) to subjects for the prevention of atherosclerosis or its signs, symptoms, or complications. In some embodiments, subjects with a predisposition to atherosclerosis may exhibit one or more of the following characteristics: advanced age, family history of heart disease, biological condition, and high blood cholesterol. In some embodiments, the biological condition includes high levels of low-density lipoprotein cholesterol (LDL-C) in the blood, low levels of high-density lipoprotein cholesterol (HDL-C) in the blood, hypertension, insulin resistance, diabetes, excess body weight, obesity, sleep apnea, contributing lifestyle choices, and / or contributing behavioral habits. In some embodiments, behavioral habits include smoking and / or alcohol consumption. In some embodiments, lifestyle choices include an inactive lifestyle and / or high stress levels.

[0267] In exemplary embodiments, the present disclosure provides administration of the CD dimer of this disclosure in combination with one or more optional additional agonists to a patient suffering from atherosclerosis. The patient may exhibit one or more signs or symptoms of atherosclerosis. Atherosclerosis can be diagnosed based on one or more of the following: Doppler ultrasound, ankle-brachial index, electrocardiogram, stress test, angiography (optionally using cardiac catheterization), computed tomography (CT), magnetic resonance angiography (MRA), or other arterial imaging or blood flow measurement methods.

[0268] In exemplary embodiments, the present disclosure provides administration in combination therapy comprising the CD dimer and one or more additional therapies. These combination therapies for treating atherosclerosis may include the CD dimer of the present disclosure and other therapies for the treatment or prevention of atherosclerosis, such as anticholesterol agents, antihypertensive agents, antiplatelet agents, nutritional supplements, or surgery or behavioral interventions, including, but not limited to, those described below. Further combination therapies may include the CD dimer of the present disclosure and other therapies for the treatment of heart failure, such as one or more or a combination thereof, of aldosterone antagonists, ACE inhibitors, ARBs (angiotensin II receptor blockers), ARNIs (angiotensin receptor neprilysin inhibitors), beta-blockers, vasodilators, calcium channel blockers, digoxin, diuretics, cardiac pump drugs, potassium, magnesium, selective sinoatrial node inhibitors. Combination therapies for treating atrophic age-related macular degeneration (AMD) or Stargardt disease include the CD dimer of this disclosure and other therapies for treating AMD, such as AREDS nutritional supplements containing LBS-008 (Belite Bio) (a non-retinoid antagonist of retinol-binding protein 4), vitamin C and vitamin E, beta-carotene, zinc, and copper; AREDS2 nutritional supplements containing vitamin C and vitamin E, zinc, copper, lutein, zeaxanthin, and omega-3 fatty acids; or combinations thereof. Combination therapies for treating Alzheimer's disease include the CD dimer of this disclosure and one or more cholinesterase inhibitors (ARICEPT(R), EXELON(R), RAZADYNE(R)) and memantine (NAMENDA(R)), or combinations thereof. Combination therapy for Niemann-Pick disease comprises the CD dimer of this disclosure and one or more of the following: miglustat (ZAVESCA(R)), HPβCD (TRAPPSOL CYCLO, VTS-270), and physiotherapy. This combination therapy may be administered simultaneously, essentially simultaneously, or sequentially in any of these orders.Combination therapy may be administered simultaneously with a single formulation, separately, or optionally using a dosing kit or pack containing a combination of each drug, which may be a pre-measured, convenient format providing, for example, one or more single doses of each drug in combination. Combination therapy may exhibit synergistic effects, with the combined therapy exceeding the effects of individual monotherapy. Combination therapy generally involves administering effective doses of the C. della bicarbonate dimer and the combined therapy, but combination therapy can yield effective treatment with lower doses of C. della bicarbonate and / or the combined therapy, which may advantageously reduce the side effects associated with the usual (non-combination) doses.

[0269] Combination therapies may include therapies for the treatment or prevention of diseases or conditions associated with atherosclerosis, such as coronary artery disease, angina pectoris, heart attack, cerebrovascular disease, transient ischemic attack, and / or peripheral artery disease. Combination therapies may also include therapies for the treatment or prevention of conditions that may contribute to the formation of atherosclerosis and / or worsening of prognosis, such as hypertension, hypercholesterolemia, hyperglycemia, and diabetes.

[0270] In exemplary embodiments, the CD dimer of the present invention is an anticholesterol agent, for example, a fibrate or statin agent, for example, ADVICOR(R) (niacin sustained-release / lovastatin), ALTOPREV(R) (lovastatin sustained-release), CADUET(R) (amlodipine / atorvastatin combination), CRESTOR(R) (rosuvastatin), JUVISYNC(R) (sitagliptin / simvastatin), LESCOL(R) (fluvastatin), LESCOL It is administered concurrently with XL (fluvastatin sustained-release), LIPITOR(R) (atorvastatin), LIVALO(R) (pitavastatin), MEVACOR(R) (lovastatin), PRAVACHOL(R) (pravastatin), SIMCOR(R) (niacin sustained-release / simvastatin), VYTORIN(R) (ezetimibe / simvastatin), and / or ZOCOR(R) (simvastatin), etc. Anticholesterol agents may be administered in doses effective in preventing or treating hypercholesterolemia.

[0271] In exemplary embodiments, the CD dimer of the present invention is administered concurrently with an antiplatelet drug, such as aspirin.

[0272] In exemplary embodiments, the CD dimer of the present invention is administered concurrently with an antihypertensive drug. Examples of antihypertensive drugs include beta-blockers, angiotensin-converting enzyme (ACE) inhibitors, calcium channel blockers, and / or diuretics.

[0273] In exemplary embodiments, the CD dimer of the present invention is administered concurrently with one or more nutritional supplements, such as alpha-linolenic acid (ALA), barley, beta-sitosterol, black tea, psyllium husk, calcium, cocoa, cod liver oil, coenzyme Q10, fish oil, folic acid, garlic, green tea, niacin, oat bran, omega-3 fatty acids (eicosapentaenoic acid (EPA) and / or docosahexaenoic acid (DHA), etc.), cytostanol, and / or vitamin C.

[0274] Exemplary combination therapies include counseling and / or support for interventions in the patient's behavior and / or lifestyle, such as smoking cessation, exercise, and healthy eating, such as diets low in low-density lipoprotein (LDL) and optionally high in high-density lipoprotein (HDL).

[0275] Exemplary combination therapies include surgical interventions, such as angioplasty, stent placement, or both.

[0276] The methods of the present invention are useful for treating or preventing atherosclerosis in human subjects. In some cases, patients are healthy except for exhibiting atherosclerosis. For example, a patient may not exhibit any other risk factors for cardiovascular, thrombotic, or other diseases or disorders at the time of treatment. However, in other cases, patients are selected based on having been diagnosed with or being at risk of developing a disease or disorder caused by or associated with atherosclerosis. For example, at the time of or before administration of the pharmaceutical composition of the present invention, a patient may have been diagnosed with or been identified as being at risk of developing a cardiovascular disease or disorder, such as coronary artery disease, acute myocardial infarction, asymptomatic carotid atherosclerosis, stroke, peripheral artery occlusive disease, etc. The cardiovascular disease or disorder may, in some cases, be hypercholesterolemia.

[0277] In other cases, at the time of administration of the pharmaceutical composition of the present invention, or prior to administration, the patient may have been diagnosed with or identified as being at risk of developing atherosclerosis.

[0278] In other cases, patients to be treated by the method of the present invention are selected based on one or more factors selected from the group consisting of age (e.g., older than 40, 45, 50, 55, 60, 65, 70, 75, or 80 years of age), race, sex (male or female), exercise habits (e.g., regularly exercises, does not exercise), other pre-existing medical conditions (e.g., type II diabetes, hypertension, etc.), and current medical status (e.g., currently taking statin drugs, e.g., cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin, pravastatin, etc., beta-blockers, niacin, etc.).

[0279] Embodiments of the present invention provide compositions and methods for treating or preventing atherosclerosis and other age-related diseases. 7KC is the most abundant non-enzymatically produced oxysterol in atherosclerotic plaques and is thought to be a contributing factor to the development of atherosclerosis. Treatment using the CD dimer of the present invention is expected to be beneficial in preventing and / or improving atherosclerotic plaque formation.

[0280] Embodiments of the present invention provide compositions and methods for the treatment or prevention of diseases and conditions in which 7KC is involved. These include, but are not limited to, age-related diseases such as age-related macular degeneration (AMD), Alzheimer's disease, and lysosomal storage disorders such as Niemann-Pick type C (NPC). 7KC is also involved in the pathogenesis of cystic fibrosis, liver injury and failure, and hypercholesterolemia. The increased rate of dementia in hypercholesterolemia is associated with 7KC accumulation. [Examples]

[0281] V. Examples

[0282] Example 1. MD simulation of βCD and βCD dimers

[0283] Previously, the inventors performed various simulations of monomeric (Figures 3E-F) and dimerized (Figures 4A-H) βCD molecules that form complexes with 7KC and cholesterol. These studies ultimately demonstrated that dimerization significantly improves complex formation with these ligands. Subsequently, these types of simulations were expanded to include other types of dimers, as supported by wet lab and NMR data shown in Figures 5-6. Performing computer analysis before chemical synthesis and wet lab studies allows for theoretical testing of these molecules before physically synthesizing and testing them. A brief summary of the previous research is as follows:

[0284] Compared to monomers (Figures 3E-3F), our novel butyl-linked DS5 hydroxypropyl β-CD dimer (Figure 4A) showed significantly improved stable complex formation with 7KC and cholesterol in the GROMOS force field. The contrast between these trajectory plots and the plots for monomeric HPβCD(DS5) and unmodified βCD provides clear evidence that the dimerized version consistently and remarkably reliably binds to sterols, whether hydroxypropylated or not, compared to its corresponding monomeric version. This is consistent with our experimental data comparing monomers and dimers (Figures 5A-5Q and 6A-6D). The angles, distances, and energies are all significantly more stable than those in monomeric simulations, likely due to a more dissolved configuration. The inventors have found that when the complex is fully formed in the downward direction, the distance between the center of gravity of the ligand and the CD is less than 5 angstroms, whereas the monomer in the GROMOS force field consistently showed that the intermolecular distance exceeds 5–10 angstroms when the complex is formed. This indicates that the dimer forms a very potent and stable complex with both ligands, especially in the downward direction, and particularly compared to monomeric βCD. The data show that dimerized HPβCD has a much stronger sterol affinity overall than the monomer, and that dimerized HPβCD has a preference for 7KC because 7KC associates with at least one of the CDs for significantly longer than cholesterol.

[0285] Based on these and other early HPβCD simulations, it was concluded that the GROMOS force field at the ideal inclusion complex initiation position (both directions) yields the best and most dynamic results for these complexes. This lengthy initial analysis was crucial in establishing precedents for modeling these novel molecules, thereby enabling faster and more targeted simulations for other dimers. Therefore, the molecular dynamics analysis has been extended using various promising linkers and substitutions, including triazole and butyl-linked methyl βCD, sulfobutyl βCD, and quaternary ammonium βCD, all of which are DS4 (Figures 4A-4H). Using a triazole linker with HPβCD dimers appeared to produce less stable complexes, but these complexes were more specific to 7KC than to cholesterol (Figure 4B). For methyl dimers (Figures 4C-4D), those with a butyl linker formed the most stable complex, and both linker cases appeared to favor the upward direction, although the interactions were very similar for the two methyl dimers tested. Regarding methyl substitution, it is difficult to distinguish which is actually more effective, but both types of linkers readily form complexes with both ligands. From the trajectory, it became clear that the 7KC head group is not fully occupying the dimer cavity, but is stably maintained between the two sister monomers.

[0286] Negatively charged sulfobutyl dimers (Figures 4E-4F) exhibit a similar pattern to methyl and hydroxypropyl dimers, in which case the triazole linker forms a slightly unstable complex, but enables 7KC specificity. The charged, bulky sulfobutyl group appears to interact very favorably with both 7KC and cholesterol, but in both linker cases, the only separated complex is that of cholesterol. This suggests that sulfobutyl dimers likely have very good specificity for 7KC compared to methyl and hydroxypropyl dimers.

[0287] To further investigate the potential of charged substituents, MD analysis (Figures 4G-4H) was performed on positively charged quaternary ammonium βCD of DS4. The QA dimer exhibited similar properties to the SB dimer, with better 7KC specificity as seen with the triazole linker, but overall complex formation with both ligands was good. A particularly strong downward complex was formed between butyl-linked QAβCD and cholesterol, but triazole-linked QAβCD formed the most stable complex with downward 7KC.

[0288] Based on the predictive nature of these simulations, we extend this type of analysis to include the CD dimers described here.

[0289] Example 2. MD simulation of a C6-butyl-substituted DS6 triazole-linked βCD dimer complex with sterols.

[0290] Figure 11A shows the trajectory results from MD simulations of the C6-butyl-substituted DS6 βCD dimer, which forms complexes independently with 7KC and cholesterol in both the upward and downward directions. The upper graph shows the distance between the center of gravity of the ring of O4 atoms in one of the CD monomers of the dimer and the center of gravity of the sterol over a period of 100 ns. The middle graph shows the angle formed between the principal axis of the sterol and the axis perpendicular to the ring of O4 atoms (shown in Figure 3C) over a period of 100 ns. The lower graph shows the interaction energy between the CD dimer and the sterol (i.e., the respective host and guest in the host-guest complex) over a period of 100 ns. The sterol and its direction are represented by the following colors: 7KC-up (red), 7KC-down (blue), cholesterol-up (black), and cholesterol-down (orange).

[0291] The data shows little variation between each simulation run of a given complex, indicating that the dimer forms relatively stable host-guest interactions with both 7KC and cholesterol, regardless of whether the interaction is upward or downward, and that this dimer is an effective encapsulation agent for sterol-like molecules.

[0292] Example 3. MD simulation of the C62-hydroxypropyl DS6 triazole-linked βCD dimer complex with sterols.

[0293] Figure 11B shows the trajectory results from MD simulations of the C6 2-hydroxypropyl DS6 βCD dimer, which forms complexes independently with 7KC and cholesterol in both the upward and downward directions. The upper graph shows the distance between the center of gravity of the ring of O4 atoms in one of the CD monomers of the dimer and the center of gravity of the sterol over a period of 100 ns. The middle graph shows the angle formed between the principal axis of the sterol and the axis perpendicular to the ring of O4 atoms (shown in Figure 3C) over a period of 100 ns. The lower graph shows the interaction energy between the CD dimer and the sterol (i.e., the respective host and guest in the host-guest complex) over a period of 100 ns.

[0294] The data shows little variation between each simulation run of the specific complex, indicating that the dimer forms relatively stable host-guest interactions with both 7KC and cholesterol, regardless of whether they are upward or downward, and that this dimer is an effective encapsulation agent for sterol-like molecules.

[0295] molecular dynamics methods

[0296] To gain a more comprehensive understanding of the role of CD-sterol complex formation and the dimerization of these molecules, various MD simulations were performed using GROMACS software. These simulations clearly outline the behavior of water molecules around the structure, as well as the internal dynamics of the various molecular groups within the complex. Both challenges have recently been reported to be highly specific to these types of structures. GROMACS parameters were obtained by combining the unique topology of unmodified CD [J.Phys.Chem B,118,2014,699958] with parameterization of various groups obtained from the ATB server, and were sequentially validated using reference components of known molecules and intrinsic parameters of the force field. Two inclusion complex structures, named "upper" and "lower," were prepared for the force field and ligand respectively, so as to be opposite to the orientation of the sterol molecules within the CD cavity, i.e., parallel and antiparallel to the axis of symmetry of CD aligned with their principal axes (Figure 3A). The resulting structures were solvated with approximately 3000 water molecules. During minimization, the obtained systems were adopted as the initial structures for MD simulations. An unlimited production trajectory of 100 ns was generated using a 2 fs integration time step. In all simulations, pressure and temperature were controlled at 1 bar and 298 K, respectively, using an isotropic Parrinello-Rahman barostat [J.Appl.Phys.52,1981,7182] and a velocity rescaling (V-rescale) thermostat [J.Phys.Chem.126,2007,014101]. The LINCS algorithm [J.Comput.Chem.18,1997,1463] was employed to eliminate coupled oscillations. In all cases, long-range electrostatics were handled using the particle mesh Ewald method coupled to periodic boundary conditions with a 0.15 nm grid spacing and a 1.2 nm direct spatial cutoff. Van der Waals interactions were computer-calculated using a 1.2 nm spherical cutoff. Simulation analysis was performed using standard GROMACS tools and processed using Python scripts.

[0297] Figure 3C illustrates how useful “angle” measurements are in determining how well a ligand is shielded from surrounding water molecules: zero or 180 degrees indicates the ligand is perpendicular to the plane of the CD, and therefore the two molecules are most likely to form a soluble complex, while 90 degrees indicates the ligand is parallel to the CD plane and is less likely to form a complex within the cavity. For our complex, approximately 30 degrees corresponds to the “upward” direction of complex formation (sterol head associated with the secondary plane of the CD, tail on the primary plane, and the entire ligand inserted into the cavity of the CD), and approximately 150 degrees corresponds to the “downward” direction of complex formation (sterol tail associated with the secondary plane of the CD, head on the primary plane, and the entire ligand inserted into the cavity of the CD).

[0298] Example 4. Solubilization of compounds with βCD monomer

[0299] Example 4 illustrates the ability of various substituted β-CD monomers to solubilize cholesterol and 7KC (Figures 5A-5D). Lower turbidity indicates a higher ability to solubilize a given sterol. The ability to solubilize 7KC and cholesterol decreased as the degree of substitution increased (Figures 5A and 5B). HPβCDs with lower DS showed a preferential solubilization of 7KC over cholesterol, suggesting that such HPβCDs possess specificity for 7KC. While not intended to be theoretically limiting, a possible explanation is that the maximum number of hydroxyl groups are available to form hydrogen bonds with the keto group at position 7 of 7KC. However, this theory is not necessary for carrying out the present invention.

[0300] Example 5. Synthesis of HPβCD-substituted CD dimers

[0301] Figures 7C and 7H show the specific molecules synthesized in this embodiment.

[0302] This example describes the synthesis of substituted CD dimers, first linked with a butyl linker, and then linked with a triazole-containing linker.

[0303] For DS measurements, 1H and 2D NMR spectra are recorded at 600 MHz using a Varian VXR-600, with the residual solvent signal used as an internal reference. To elucidate the structure, the sample is dissolved in DMSO-d6 / D2O. The FID signal is recorded after scanning at least 16 times to obtain a spectral window containing at least 0 ppm to +10 ppm. The average DS can be calculated by setting the integral value of the anomeric region to 14 (where 14 is the number of anomeric protons in the beta-CD dimer) and dividing the integral value of the alkyl region by 3 (see Figure 7D).

[0304] General explanation of synthesis and characterization

[0305] HP(βCD-butyl-βCD) homodimer

[0306] The synthesis of HPβCD butyl-linked dimers was achieved through a three-step synthesis (see Figure 7A). The starting material was monomeric β-CD with a tert-butyldimethylsilyl group protected on the primary side (TBDMS-βCD, CycloLab, Budapest, Hungary).

[0307] Secondary dimerization was achieved using TBDMS-βCD, under anhydrous conditions, and sodium hydride as the base. A dialkylating agent was added dropwise to the heterogeneous reaction mixture and the reaction was allowed to proceed completely at room temperature.

[0308] The βCD dimer (TBDMS-βCD-butyl-βCD-TBDMS), with the primary side protected, was purified by chromatography using isocratic elution (chloroform:methanol:water = 50:8:0.8 (v / v / v) as the eluent). The identity of the product was confirmed by MALDI and NMR analysis of the compound.

[0309] Desilylation (deprotection) was performed in THF at room temperature using tetrabutylammonium fluoride. The βCD dimer (βCD-butyl-βCD) was purified by chromatography using isocratic elution (eluent: 1,4-dioxane:25% NH3 (aqueous solution) = 10:7 (v / v)). The identity of the product was confirmed by MALDI and TLC analysis of the compound.

[0310] Hydroxypropylation of the βCD dimer was achieved under aqueous conditions at room temperature using sodium hydroxide as the base. Purification of the hydroxypropylated βCD dimer (HP(βCD-butyl-βCD)) dimer was based on ion exchange resin treatment, activated carbon clarification, and large-scale dialysis. The identity and structure of the product were confirmed by MALDI and NMR analysis of the compound (Figures 7B, 7D, and 7E). The HSQC is a type of HSQC that can distinguish between CH3 (methyl) / CH (methine) correlations (visualized as red crossover peaks, such as C1, C2, C3, C4, and C5 of the cyclodextrin unit, or alternatively, as the methyl group in the case of methyl-substituted CD, for example) and CH2 correlations (visualized as a blue crossover peak, such as the C6 signal of the cyclodextrin unit), because they are in opposite phases. These colors may not be distinguishable in black and white reproductions, but analysis of the original color HSQC diagram reveals the expected features, including the crossover peaks of red and blue.

[0311] HP(αCD-butyl-αCD) homodimer

[0312] The synthesis of butyl-linked HPαCD dimers is achieved through a three-step synthesis (see Figure 12A). The starting material is monomeric αCD with the primary side protected by a tert-butyldimethylsilyl group (TBDMS-αCD, CycloLab, Budapest, Hungary).

[0313] Secondary dimerization was achieved using TBDMS-αCD, under anhydrous conditions, and sodium hydride as the base. A dialkylating agent was added dropwise to the heterogeneous reaction mixture and allowed to react completely at room temperature.

[0314] The αCD dimer (TBDMS-αCD-butyl-αCD-TBDMS) with the primary side protected is purified by chromatography using isocratic elution (chloroform:methanol:water = 50:8:0.8 (v / v / v) as the eluent).

[0315] Desilylation (deprotection) is performed in THF at room temperature using tetrabutylammonium fluoride. The αCD dimer (αCD-butyl-αCD) is purified by chromatography using isocratic elution (eluent: 1,4-dioxane:25% NH3 aqueous solution = 10:7 (v / v)). Hydroxypropylation of the αCD-butyl-αCD dimer is achieved under aqueous conditions at room temperature using sodium hydroxide as the base. Purification of the hydroxypropylated αCD dimer (HP(αCD-butyl-αCD)) is based on ion exchange resin treatment, activated carbon clarification, and large-scale dialysis.

[0316] HP(βCD-triazole-βCD) homodimer

[0317] The synthesis of hydroxypropylated β-CD dimers linked via a secondary surface at a single triazole moiety is carried out in four steps (Figure 7F). The first step is the preparation of an azido linker (3-azido-1-bromo-propane), as this reagent is not commercially available. The second step is the construction of two βCD monomers, 2-O-monopropargyl-β-CD and 2-O-mono(3-azidopropyl)-βCD, respectively. The third step of the synthesis is the formation of a βCD-triazole-βCD dimer nucleus by copper-catalyzed azido-alkyne cycloaddition, and the final step is the preparation of a series of 2-hydroxypropylated triazole linked dimers by a classical alkylation approach.

[0318] In detail, the preparation of the azid linker can be achieved by strictly limiting the amount of sodium azide and extending the addition time of the limiting reagent. The azid linker is characterized by NMR spectroscopy and TLC.

[0319] The synthesis of the two monomers is achieved by using lithium hydride as a base for the selective deprotonation of the secondary hydroxyl group. Specifically, this approach largely reacts only the hydroxyl group located at C2. As a result, the monomers prepared by this method are preferentially substituted with O2 (a single isomer). The two monomers are characterized by NMR spectroscopy, MALDI, and TLC.

[0320] Next, using copper bromide as a catalyst, the two monomers are reacted in an aqueous DMF solution to prepare a dimeric nucleus. The resulting compound, a single isomer (BCD-triazole-BCD, DS=0), is characterized by NMR spectroscopy and MALDI.

[0321] Hydroxypropylation of BCD-triazole-BCD was achieved using propylene oxide and alkaline aqueous conditions. The series of hydroxypropylated compounds were characterized by NMR spectroscopy (Figures 7I-7J) and MALDI (Figure 7G).

[0322] HPβCD-triazole-βCD (random substitution) asymmetric dimer

[0323] The synthesis of 2-hydroxypropylated βCD asymmetric dimers, linked by random substitution via a secondary plane at a single triazole moiety, is carried out in three steps (Figures 20A-20F). The first step is the preparation of an azido linker (3-azido-1-bromo-propane), as this reagent is not commercially available. The second step is the construction of two βCD monomers, 2-O-mono(3-azidopropyl)-βCD and the randomly substituted asymmetric monomer, (2-hydroxypropylated)-2-O-monopropargyl-βCD. The third step of the synthesis is the formation of the final dimer by copper-catalyzed azido-alkyne cycloaddition. In the preparation of the asymmetric dimer, it is essential to customize the asymmetric monomer before cycloaddition, as the "asymmetry" of the final dimer can only be introduced at this development stage.

[0324] In detail, the preparation of the azid linker can be achieved by strictly limiting the amount of sodium azide and extending the addition time of the limiting reagent. The azid linker is characterized by NMR spectroscopy and TLC.

[0325] The monomer synthesis is achieved by using lithium hydride as the base for selective deprotonation on the secondary side. In detail, this approach causes the hydroxyl group located at C2 to react almost entirely. As a result, the monomer prepared by this method is mainly substituted with O2. 2-O-mono(3-azidopropyl)-βCD is prepared in one step according to the method described above and obtained as a single isomer. To introduce "asymmetry" to the second monomer, 2-O-monopropargyl-βCD is 2-hydroxypropylated using propylene oxide and alkaline aqueous solution conditions. The two monomers are characterized by NMR spectroscopy, MALDI, and TLC.

[0326] Next, using copper bromide as a catalyst, the two monomers are reacted in an aqueous DMF solution to prepare an asymmetric dimer. The resulting compound, HPβCD'-triazole-βCD DS3 (random substitution) asymmetric dimer, is characterized by NMR spectroscopy and MALDI.

[0327] C6HPβCD-triazole-βCD DS3 asymmetric dimer

[0328] The synthesis of the C6-primary (2-hydroxypropylated)-βCD asymmetric dimer DS3, linked via a secondary surface by a single triazole moiety, is carried out in three steps (Figures 20B-20F). The first step is the preparation of the azido linker (3-azido-1-bromopropane) and the protected 2-hydroxypropylating agent, as these reagents are not commercially available (Figure 20C).

[0329] The second step involves constructing the two βCD monomers, 2-O-mono(3-azidopropyl)-βCD and the asymmetric monomer tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD (Figures 20D-20E).

[0330] The third step in the synthesis is the formation of the final dimer by copper-catalyzed azide-alkyne cycloaddition (Figure 20F).

[0331] In the preparation of asymmetric dimers, the "asymmetry" of the final dimer can only be introduced at this development stage; therefore, it is essential to customize the asymmetric monomer before cycloaddition.

[0332] In detail, the preparation of the azid linker can be achieved by strictly limiting the amount of sodium azide and extending the addition time of the limiting reagent. The azid linker is characterized by NMR spectroscopy and TLC. The synthesis of the protected 2-hydroxypropylating agent (1-bromo-2-benzyloxy-propane) is achieved in two steps. Propylene oxide is reacted with benzyl alcohol as a solvent under acidic conditions to obtain 2-benzyloxy-1-propanol. The resulting alcohol is then converted to a bromo analog using potassium bromide in acetonitrile under acidic conditions.

[0333] The monomer synthesis is achieved by using lithium hydride as a base for selective deprotonation of the secondary side. In detail, this approach causes the hydroxyl group located at C2 to react almost entirely. As a result, the monomer prepared by this method is mainly substituted with O2. 2-O-mono(3-azidopropyl)-βCD is prepared in one step according to the method described above and obtained as a single isomer. To introduce "asymmetry" only on the primary side of the second monomer, 2-O-monopropargyl-βCD is modified according to a multi-step synthetic procedure (Figure 20C). 2-O-monopropargyl-βCD is selectively protected on the primary side with a tert-butyldimethylsilyl moiety, and then the secondary side is comprehensively modified with benzyl bromide under phase-transfer catalytic (PTC) conditions to produce an asymmetrically protected monomer, per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD. Selective removal of the tert-butyldimethylsilyl moiety is readily achieved with tetrabutylammonium fluoride in THF. The key intermediate, per-2,3-O-benzyl-2-O-monopropargyl-βCD, is reacted with a protected 2-hydroxypropylating agent (1-bromo-2-benzyloxypropane) under PTC conditions. The desired DS3 is achieved by strictly limiting the amount of base (KOH). Finally, comprehensive deprotection of the benzyl group by hydrazine-mediated transfer hydrogenation yields the asymmetric monomer, tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD. The two monomers are characterized by NMR spectroscopy, MALDI, and TLC.

[0334] Next, using copper bromide as a catalyst, the two monomers are reacted in an aqueous DMF solution to prepare an asymmetric dimer. The resulting compound, C6HPβCD'-triazole-βCD DS3 asymmetric dimer, is characterized by NMR spectroscopy and MALDI.

[0335] C6HPβCD-triazole-βCD DS7 asymmetric dimer

[0336] The synthesis of the C6-primary (2-hydroxypropylated)-βCD asymmetric dimer DS7, which is linked via a secondary surface by a single triazole moiety and completely substituted, is carried out in a three-step procedure (Figures 21A-21D). The first step is the preparation of the azido linker (3-azido-1-bromo-propane) and the protected version of the 2-hydroxypropylating agent, as these reagents are not commercially available (Figure 21A).

[0337] The second step involves constructing two βCD monomers, 2-O-mono(3-azidopropyl)-βCD and the asymmetric monomer per-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD (Figures 21B-21C).

[0338] The third step in the synthesis is the formation of the final dimer by copper-catalyzed azide-alkyne cycloaddition (Figure 21D).

[0339] In the preparation of asymmetric dimers, the "asymmetry" of the final dimer can only be introduced at this development stage; therefore, it is essential to customize the asymmetric monomer before cycloaddition.

[0340] In detail, the preparation of the azid linker can be achieved by strictly limiting the amount of sodium azide and extending the addition time of the limiting reagent. The azid linker is characterized by NMR spectroscopy and TLC. The synthesis of the protected 2-hydroxypropylating agent (1-bromo-2-benzyloxy-propane) is achieved in two steps. Propylene oxide is reacted with benzyl alcohol as a solvent under acidic conditions to obtain 2-benzyloxy-1-propanol. The resulting alcohol is then converted to a bromo analog using potassium bromide in acetonitrile under acidic conditions.

[0341] The monomer synthesis is achieved by using lithium hydride as a base for selective deprotonation of the secondary side. In detail, this approach causes the hydroxyl group located at C2 to react almost entirely. As a result, the monomer prepared by this method is mainly substituted with O2. 2-O-mono(3-azidopropyl)-βCD is prepared in one step according to the method described above and obtained as a single isomer. To introduce "asymmetry" only on the primary side of the second monomer, 2-O-monopropargyl-βCD is modified by a multi-step synthetic procedure (Figure 21B). 2-O-monopropargyl-βCD is modified by selectively protecting the primary side with a tert-butyldimethylsilyl moiety, followed by comprehensive modification of the secondary side with benzyl bromide under phase-transfer catalytic (PTC) conditions, thereby producing an asymmetrically protected monomer, per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD. Selective removal of the tert-butyldimethylsilyl moiety is readily achieved with tetrabutylammonium fluoride in THF. The key intermediate, per-2,3-O-benzyl-2-O-monopropargyl-βCD, is reacted with a protected excess of a 2-hydroxypropylating agent (1-bromo-2-benzyloxypropane) under PTC conditions. The desired DS7, a complete substitution of the primary side, is achieved by simultaneously using an excess of a base (KOH) and a protected excess of the 2-hydroxypropylating agent. Finally, comprehensive deprotection of the benzyl group by hydrazine-mediated transfer hydrogenation yields the asymmetric monomer, per-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD. The two monomers are characterized by NMR spectroscopy, MALDI, and TLC.

[0342] Next, using copper bromide as a catalyst, the two monomers are reacted in an aqueous DMF solution to prepare an asymmetric dimer. The resulting compound, C6HPβCD'-triazole-βCD DS7 asymmetric dimer, is characterized by NMR spectroscopy and MALDI.

[0343] HP(αCD-triazole-αCD) homodimer

[0344] The preparation of hydroxypropylated αCD dimers linked via a secondary surface at a single triazole moiety is carried out in four steps (Figure 12B). The first step is the preparation of an azido linker (3-azido-1-bromo-propane), as this reagent is not commercially available. The second step is the preparation of two αCD monomers, 2-O-monopropargyl-αCD and 2-O-mono(3-azidopropyl)-αCD, respectively. The third step of the synthesis is the formation of an αCD-triazole-αCD dimer by copper-catalyzed azido-alkyne cycloaddition, and the final step is the preparation of a series of 2-hydroxypropylated triazole linked dimers by a classical alkylation approach.

[0345] In detail, the preparation of the azid linker can be achieved by strictly limiting the amount of sodium azide and extending the addition time of the limiting reagent. The azid linker is then characterized by NMR spectroscopy and TLC.

[0346] The synthesis of the two monomers is achieved by using lithium hydride as the base for selective deprotonation on the secondary side. Specifically, this approach causes the hydroxyl group located at C2 to react almost entirely. As a result, the monomers prepared by this method are mainly substituted with O2 (a single isomer).

[0347] Next, the αCD-triazole-αCD dimer is prepared by reacting two monomers.

[0348] Hydroxypropylation of the αCD-triazole-αCD dimer was achieved using propylene oxide and alkaline aqueous conditions.

[0349] HP(αCD-butyl-βCD) heterodimer

[0350] The preparation of butyl-linked HPαCD-βCD dimers was achieved through a three-step synthesis (see Figure 16A). The starting materials were monomeric αCD and βCD with the primary side protected by a tert-butyldimethylsilyl group (TBDMS-αCD and TBDMS-βCD, CycloLab, Budapest, Hungary).

[0351] Secondary dimerization was achieved by using TBDMS-αCD and TBDMS-βCD, under anhydrous conditions, and equimolar amounts of sodium hydride as the base. A dialkylating agent was added dropwise to the heterogeneous reaction mixture and the reaction was allowed to proceed completely at room temperature.

[0352] The αCD-βCD dimer (TBDMS-αCD-butyl-βCD-TBDMS), with the primary side protected, was purified by chromatography using isocratic elution (chloroform:methanol:water = 50:8:0.8 (v / v / v) as the eluent).

[0353] Desilylation (deprotection) was performed in THF at room temperature using tetrabutylammonium fluoride. The αCD-βCD dimer (αCD-butyl-βCD) was purified by chromatography using isocratic elution (eluent: 1,4-dioxane:25% NH3 aqueous solution = 10:7 (v / v)).

[0354] Hydroxypropylation of the αCD-butyl-βCD dimer was achieved under aqueous conditions at room temperature using sodium hydroxide as the base. Purification of the hydroxypropylated αCD-βCD dimer (HP(αCD-butyl-βCD)) was based on ion exchange resin treatment, activated carbon clarification, and large-scale dialysis.

[0355] HP(αCD-triazole-βCD) heterodimer and HP(βCD-triazole-αCD) heterodimer

[0356] The preparation of hydroxypropylated αCD-βCD dimers and hydroxypropylated βCD-αCD dimers linked via a secondary surface by a single triazole moiety is carried out in four steps (Figure 16B). The first step is the preparation of an azido linker (3-azido-1-bromo-propane), as this reagent is not commercially available. The second step is the preparation of four monomers: 2-O-monopropargyl-αCD, 2-O-monopropargyl-βCD, and 2-O-mono(3-azidopropyl)-αCD and 2-O-mono(3-azidopropyl)-βCD. The third step of the synthesis is the formation of two dimer nuclei, αCD-triazole-βCD dimer and βCD-triazole-αCD dimer, by copper-catalyzed azido-alkyne cycloaddition, and the final step is the preparation of a series of 2-hydroxypropylated triazole linked dimers by a classical alkylation approach.

[0357] In detail, the preparation of the azid linker can be achieved by strictly limiting the amount of sodium azide and extending the addition time of the limiting reagent. The azid linker is then characterized by NMR spectroscopy and TLC.

[0358] The synthesis of the four monomers is achieved by using lithium hydride as the base for selective deprotonation on the secondary side. Specifically, this approach causes the hydroxyl group located at C2 to react almost entirely. As a result, the monomers prepared by this method are mainly substituted with O2 (a single isomer).

[0359] The preparation of the two dimeric nuclei, the αCD-triazole-βCD dimer and the βCD-triazole-αCD dimer, is achieved by reacting the 2-O-monopropargyl-αCD monomer with the 2-O-mono(3-azidopropyl)-βCD monomer, and by reacting the 2-O-monopropargyl-βCD monomer with the 2-O-mono(3-azidopropyl)-αCD monomer, respectively.

[0360] Hydroxypropylation of αCD-triazole-βCD dimers and βCD-triazole-αCD dimers is achieved using propylene oxide and alkaline aqueous conditions.

[0361] Detailed explanation of the synthesis of (HP(βCD-butyl-βCD) homodimer)

[0362] Step 1: Secondary dimerization of TBDMS-βCD

[0363] Under an inert atmosphere, anhydrous TBDMS-βCD (10 g, 5.17 mmol) was dissolved in THF (400 mL), and sodium hydride (2.5 g, 50 mmol) was carefully added little by little (over 30 minutes). Upon addition of sodium hydride, hydrogen was formed, and the suspension foamed vigorously. After stirring for 15 minutes, the reaction mixture gelled (became viscous), and stirring became difficult. To break up the gel, the reaction mixture was heated until gentle reflux occurred, and reflux was maintained for 30 minutes. The yellowish heterogeneous suspension became easier to stir, and the gel-like substance disappeared. The reaction mixture was cooled to room temperature in a water bath. When the alkylating agent 1,4-dibromobutane (1.25 mL, 2.25 g, 10.5 mmol) was added dropwise (over 15 minutes), the color of the reaction mixture turned dark orange.

[0364] The brownish suspension was stirred overnight under an inert atmosphere. The conversion rate was estimated to be 10% to 15% by TLC (eluent:chloroform:methanol:water = 50:10:1, v / v / v), so it was determined that post-treatment was possible.

[0365] The reaction mixture was rapidly cooled with methanol (30 mL), concentrated under reduced pressure (approximately 20 mL), and precipitated with water (200 mL). The crude reaction product was filtered through a sintered glass filter and thoroughly washed with water (3 × 300 mL). The crude material was dried in a drying box in the presence of KOH and P2O5 until it reached a certain weight (recovered material: 12.1 g).

[0366] The crude reaction product was purified by chromatography, and the fraction containing the product was collected based on TLC analysis. This fraction was evaporated under reduced pressure until dry to obtain a white substance. This was dried in a drying box in the presence of KOH and P2O5 until it reached a fixed weight (TBDMS-βCD-BUT-βCD-TBDMS, 3.5 g).

[0367] Step 2: Deprotection of TBDMS-βCD butyl-linked dimer

[0368] Under an inert atmosphere, anhydrous TBDMS-βCD-BUT-βCD-TBDMS (3.5 g, 0.89 mmol) was dissolved in THF (250 mL), and tetrabutylammonium fluoride (8.75 g, 33.47 mmol) was added all at once to the yellowish solution. After stirring at room temperature for 30 minutes, the reaction mixture turned dark green. The reaction mixture was stirred at room temperature overnight. TLC analysis (1,4-dioxane:25% NH3=10:7 (v / v)) revealed that the reaction was not complete, so a second volume of tetrabutylammonium fluoride (4 g, 13.3 mmol) was added to the container. The reaction mixture was gently heated until reflux was achieved and continued for 2 hours. At this stage, the reaction transformation was complete, as no starting materials were detectable by TLC. The reaction mixture was cooled to room temperature, concentrated under reduced pressure (to approximately 10 mL), and methanol (200 mL) was added to obtain a white precipitate. The solid was filtered and analyzed by TLC, and dried in a drying box in the presence of KOH and P2O5 until a constant weight was obtained (1.2 g). TLC analysis revealed that the substance contained a negligible amount (≤3%) of tetrabutylammonium fluoride. The mother liquor was concentrated under reduced pressure (to approximately 10 mL), purified by chromatography (eluent: 1,4-dioxane:NH3=10:7v / v), and the fraction containing the product was collected and evaporated under reduced pressure until dry to obtain a white substance. This was dried in a drying box in the presence of KOH and P2O5 until a constant weight was obtained (βCD-BUT-βCD, 0.55 g).

[0369] Step 3: Hydroxypropylation of βCD-butyl-βCD homodimer

[0370] βCD-butyl-βCD DS0 (0.5 g, 0.21 mmol) was suspended in water (10 mL), and sodium hydroxide (0.1 g, 2.5 mmol) was added to the reaction vessel, resulting in a slightly yellow solution. The reaction mixture was cooled in a water bath (10 °C), and propylene oxide (0.5 mL, 0.415 g, 7.14 mmol) was added all at once. The reaction vessel was flushed with argon, sealed, and stirred at room temperature for 2 days. The reaction mixture was concentrated under reduced pressure until a viscous syrup was obtained, and this was precipitated with acetone (50 mL). The white solid was filtered through a sintered glass filter and thoroughly washed with acetone (3 × 15 mL). This substance was dissolved in water (50 mL), treated with ion exchange resin (to remove salts), clarified with activated carbon, membrane filtered, and dialyzed against purified water for 1 day. The residue was evaporated under reduced pressure until dry to obtain a white solid (0.8 g).

[0371] Detailed explanation of the synthesis of (HP(αCD-butyl-αCD) homodimer)

[0372] Step 1: Secondary dimerization of TBDMS-αCD

[0373] Under an inert atmosphere, anhydrous TBDMS-αCD (10 g, 6.03 mmol) was dissolved in THF (400 mL), and sodium hydride (2.9 g, 58 mmol) was carefully added little by little (over 30 minutes). Upon addition of sodium hydride, hydrogen was formed, and the suspension foamed vigorously. After stirring for 15 minutes, the reaction mixture gelled (became viscous), making stirring difficult. To break up the gel, the reaction mixture was heated until gentle reflux occurred, and reflux was maintained for 30 minutes. The yellowish heterogeneous suspension became easier to stir, and the gel-like structure disappeared. The reaction mixture was cooled to room temperature in a water bath. When the alkylating agent 1,4-dibromobutane (1.45 mL, 2.61 g, 12.2 mmol) was added dropwise (over 15 minutes), the color of the reaction mixture turned dark orange.

[0374] The brownish suspension is stirred overnight under an inert atmosphere. The conversion rate is estimated to be 10% to 15% by TLC (eluent:chloroform:methanol:water = 50:10:1, v / v / v), so it is determined that post-treatment is possible.

[0375] The reaction mixture is rapidly cooled with methanol (30 mL), concentrated under reduced pressure (approximately 20 mL), and precipitated with water (200 mL). The crude reaction product is filtered through a sintered glass filter and thoroughly washed with water (3 × 300 mL). The crude material is dried in a drying box in the presence of KOH and P2O5 until it reaches a certain weight (recovered material: 11.2 g).

[0376] The crude reaction product is purified by chromatography, and the fraction containing the product is collected based on TLC analysis. This fraction is evaporated under reduced pressure until dry to obtain a white substance. This is dried in a drying box in the presence of KOH and P2O5 until it reaches a constant weight (TBDMS-αCD-butyl-αCD-TBDMS, 3.3 g).

[0377] Step 2: Deprotection of the butyl-linked TBDMS-αCD dimer

[0378] Under an inert atmosphere, anhydrous TBDMS-αCD-butyl-αCD-TBDMS dimer (3.3 g, 0.98 mmol) was dissolved in THF (250 mL), and tetrabutylammonium fluoride (9.63 g, 36.85 mmol) was added all at once to the yellowish solution. After stirring at room temperature for 30 minutes, the reaction mixture turned dark green. The reaction mixture was stirred at room temperature overnight. TLC analysis (1,4-dioxane:25% NH3 aqueous solution = 10:7 (v / v)) revealed that the reaction was not complete, so the second volume of tetrabutylammonium fluoride (4 g, 13.3 mmol) was added to the container. The reaction mixture was gently heated until reflux was achieved and continued for 2 hours. At this stage, the reaction transformation was complete, as there were no starting materials detectable by TLC. The reaction mixture was cooled to room temperature, concentrated under reduced pressure (to approximately 10 mL), and methanol (200 mL) was added to obtain a white precipitate. The solid was filtered off and analyzed by TLC, then dried in a drying box in the presence of KOH and P2O5 until a constant weight was obtained (1.1 g). The mother liquor was concentrated under reduced pressure (to approximately 10 mL), purified by chromatography (eluent: 1,4-dioxane: 25% NH3 aqueous solution = 10:7 v / v), and the fraction containing the product was collected based on TLC analysis. This was evaporated under reduced pressure until dry to obtain a white substance. This was dried in a drying box in the presence of KOH and P2O5 until a constant weight was obtained (αCD-butyl-αCD dimer, 0.52 g).

[0379] Step 3: Hydroxypropylation of αCD-butyl-αCD homodimer

[0380] Suspend αCD-butyl-αCD dimer (0.52 g, 0.26 mmol) in water (10 mL), and add sodium hydroxide (0.1 g, 2.5 mmol) to the reaction vessel. The mixture will become a slightly yellow solution. Cool the reaction mixture in a water bath (10 °C) and add propylene oxide (0.5 mL, 0.415 g, 7.14 mmol) all at once. Flash the reaction vessel with argon, seal it, and stir at room temperature for 2 days. Concentrate the reaction mixture under reduced pressure until a viscous syrup is obtained, and precipitate it with acetone (50 mL). Filter the white solid through a sintered glass filter and wash thoroughly with acetone (3 × 15 mL). Dissolve this substance in water (50 mL), treat with ion exchange resin (to remove salts), clarify with activated carbon, filter by membrane, and dialyze against purified water for 1 day. Evaporate the residue under reduced pressure until dry to obtain a white solid (0.6 g).

[0381] Detailed explanation of the synthesis of (HP(βCD-triazole-βCD) homodimer)

[0382] Step 1: Preparation of Azidrinker

[0383] While vigorously stirring, 1,3-dibromopropane (10 mL, 20.18 g, 0.1 mol) was dissolved in 40 mL of DMSO. A solution of sodium azide (6.7 g, 0.1 mol) in DMSO (240 mL) was prepared and added dropwise to the 1,2-dihalopropane solution (over 2 hours). The solution was stirred overnight at room temperature. The crude reaction product was then extracted with n-hexane (3 × 100 mL), the collected organic phase was back-extracted with water (3 × 50 mL), and the organic phase was carefully evaporated under reduced pressure (strictly at 40°C and 400 mbar; otherwise, the target compound may be removed by distillation). The oily residue was purified by chromatography (isocratic elution with n-hexane-siRNA = 98:2 as the eluent). Based on TLC analysis, appropriate fractions are collected and concentrated under reduced pressure to obtain the target compound as a viscous oily substance (which can be stored in a light-shielded refrigerated container under an inert atmosphere). The compound is visualized by immersing the TLC plate in a 10% triphenylphosphine solution in dichloromethane for approximately 15 seconds, drying the TLC plate at below 60°C, immersing the TLC in a 2% ninhydrin ethanol solution for approximately 15 seconds, and finally drying the TLC plate at below 60°C. The target compound appears as purple spots on the TLC plate.

[0384] Step 2.1: Preparation of 2-O-monopropargyl-βCD

[0385] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred under N2 at room temperature for 12-24 hours until clear. Then, propargyl bromide (1.97 mL, 17.62 mmol) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution) shows spots corresponding to 2-O-monopropargylation and nonpropargylation of βCD, respectively. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved with the minimum volume of water. Add silica gel (40 g) and remove the solvent under vacuum until a powdery residue is obtained. Place this crude mixture on the top of a silica column (25 × 6 cm) and perform chromatography (10:5:2 CH3CN-H2O-25% NH3 aqueous solution). After freeze-drying, 2-O-monopropargyl-β-CD is obtained as a solid. 2-O-propargyl-β-CD was analyzed by MALDI and NMR.

[0386] Step 2.2: Synthesis of 2-O-mono(3-azidopropyl)-βCD

[0387] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred under N2 at room temperature for 12-24 hours until clear. Then, 3-azido-1-bromopropane (3 mL) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution) shows spots corresponding to 2-O-mono(3-azidopropyl)-βCD and βCD. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture is placed on top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 aqueous solution). After drying, 2-O-mono(3-azidopropyl)-β-CD is obtained as a white solid.

[0388] Step 3: Synthesis of βCD-triazole-βCD homodimer

[0389] While stirring vigorously, suspend 2-O-monopropargyl-β-CD and 2-O-mono(3-azidopropyl)-β-CD in water (300 mL) (each at a concentration of approximately 8-12 mM). To completely dissolve the heterogeneous mixture, add approximately 300 mL of N,N-dimethylformamide (DMF) to the suspension (addition of DMF is a slightly exothermic process). Add copper bromide (2 g, 13.49 mmol) to the solution. Stir the suspension at room temperature for 1 hour. The reaction is monitored by TLC and is expected to be complete after approximately 1 hour (eluent: CH3CN:H2O:25% NH3 = 10:5:2). Filter the crude reaction product and concentrate the mother liquor under reduced pressure (60°C). Dilute the gel-like substance with water and add silica (15 g). Concentrate the heterogeneous mixture under reduced pressure until dry. This crude mixture was placed on the top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution). After drying, BCD-triazole-BCD dimers were obtained. The preparation of the BCD-triazole-BCD dimers was extensively characterized by NMR.

[0390] Step 4: HP (βCD-triazole-βCD)

[0391] A βCD-triazole-βCD dimer (1 g, 0.418 mmol), obtained by steps 1-3 above or by other methods, was suspended in water (50 mL). Sodium hydroxide (DS3 = 0.32 g, 8 mmol; DS6 = 0.74 g, 18.5 mmol; DS7 = 0.87 g, 21.75 mmol) was added to the reaction vessel, and the mixture became a slightly yellow solution. The reaction mixture was cooled in a water bath (10°C), and propylene oxide (DS3 = 0.49 mL, 0.42 g, 7.25 mmol; DS6 = 1.21 mL, 1.04 g, 17.9 mmol; DS7 = 1.46 mL, 1.7 g, 29.3 mmol) was added all at once. The reaction vessel was flushed with argon, sealed, and stirred at room temperature for 2 days. The solution was concentrated under reduced pressure until a viscous syrup was obtained, and this was precipitated with acetone (50 mL). The white solid was filtered through a sintered glass filter and thoroughly washed with acetone (3 × 15 mL). This substance was dissolved in water (50 mL), treated with ion exchange resin (to remove salts), clarified with activated carbon, filtered through membrane filtration, and dialyzed against purified water for 1 day. The residue was evaporated under reduced pressure until dry to obtain a white solid (0.8 g). The HP(βCD-triazole-βCD) dimers were analyzed by NMR (Figures 7G, ​​7I, and 7J), and their DS values ​​were calculated as shown in the figures.

[0392] Detailed explanation of the synthesis of (HP(αCD-triazole-αCD) homodimer)

[0393] Step 2.1: Preparation of 2-O-monopropargyl-αCD

[0394] To a solution of αCD (17.14 g, 17.62 mmol) in dry DMSO (400 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred at room temperature under N2 until clear (12-24 hours). Then, propargyl bromide (1.964 mL, 17.62 mmol) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% NH3 aqueous solution) shows spots corresponding to monopropargyl and nonpropargyl αCD, respectively. The solution is poured into acetone (3.5 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture was placed on top of a silica column (25 × 6 cm) and purified by chromatography (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution). After drying, 2-O-monopropargyl-αCD was obtained as a solid.

[0395] Step 2.2: Synthesis of 2-O-mono(3-azidopropyl)-αCD

[0396] To a solution of βCD (17.14 g, 17.62 mmol) in dry DMSO (400 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred under N2 at room temperature for 12-24 hours until clear. Then, 3-azido-1-bromopropane (3 mL) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution) shows spots corresponding to 2-O-mono(3-azidopropyl)-αCD and αCD. The solution is poured into acetone (3.5 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Add silica gel (40g) and remove the solvent under vacuum until a powdery residue is obtained. Place this crude mixture on the top of a silica column and perform chromatography (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution). After drying, 2-O-mono(3-azidopropyl)-αCD is obtained as a white solid.

[0397] Step 3: Synthesis of αCD-triazole-αCD dimer

[0398] While stirring vigorously, suspend 2-O-monopropargyl-αCD and 2-O-mono(3-azidopropyl)-αCD in water (300 mL) (each at a concentration of approximately 8-12 mM). To completely dissolve the heterogeneous mixture, add N,N-dimethylformamide (approximately 300 mL) to this suspension (addition of DMF is a slightly exothermic process). Add copper bromide (2 g, 13.49 mmol) to the solution. Stir the suspension at room temperature for 1 hour. Monitor the reaction by TLC and expect it to be complete after approximately 1 hour (eluent: CH3CN:H2O:NH3 = 10:5:2). Filter the crude reaction product and concentrate the mother liquor under reduced pressure (60°C). Dilute the gel-like substance with water and add silica (15 g). Concentrate the heterogeneous mixture under reduced pressure until dry. This crude mixture is placed on top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution). After drying, αCD-triazole-αCD dimer is obtained.

[0399] Step 4: HP(αCD-triazole-αCD) homodimer

[0400] The αCD-triazole-αCD dimer (1 g, 0.418 mmol) obtained by steps 1-3 above or by other methods was suspended in water (50 mL). Sodium hydroxide (DS3=0.32 g, 8 mmol; DS6=0.74 g, 18.5 mmol; DS7=0.87 g, 21.75 mmol) was added to the reaction vessel, and the mixture became a slightly yellow solution. The reaction mixture was cooled in a water bath (10°C), and propylene oxide (DS3=0.49 mL, 0.42 g, 7.25 mmol; DS6=1.21 mL, 1.04 g, 17.9 mmol; DS7=1.46 mL, 1.7 g, 29.3 mmol) was added all at once. The reaction vessel was flushed with argon, sealed, and stirred at room temperature for 2 days. The solution was concentrated under reduced pressure until a viscous syrup was obtained, and this was precipitated with acetone (50 mL). The white solid was filtered through a sintered glass filter and thoroughly washed with acetone (3 × 15 mL). This substance was dissolved in water (50 mL), treated with ion exchange resin (to remove salts), clarified with activated carbon, filtered through membrane filtration, and dialyzed against purified water for 1 day. The residue was evaporated under reduced pressure until dry to obtain a white solid (0.6 g).

[0401] Detailed explanation of the synthesis of HP(αCD-butyl-βCD) heterodimer

[0402] Step 1: Secondary dimerization of TBDMS-αCD and TBDMS-βCD

[0403] Under an inert atmosphere, dissolve anhydrous TBDMS-αCD (5g, 3.01 mmol) and TBDMS-βCD (5.8g, 3.01 mmol) in THF (400 mL), and carefully add sodium hydride (2.9g, 58 mmol) little by little (over 30 minutes). Upon addition of sodium hydride, hydrogen formation occurs, and the suspension fizzes vigorously. After stirring for 15 minutes, the reaction mixture becomes viscous and difficult to stir. To break up the gel, heat the reaction mixture until gentle reflux occurs and maintain reflux for 30 minutes. The yellowish heterogeneous suspension becomes easy to stir, and the gel-like structure disappears. Cool the reaction mixture to room temperature in a water bath. When the alkylating agent 1,4-dibromobutane (1.45 mL, 2.61 g, 12.2 mmol) is added dropwise (over 15 minutes), the color of the reaction mixture turns dark orange.

[0404] The brownish suspension is stirred overnight under an inert atmosphere. The conversion rate is estimated to be 10% to 15% by TLC (eluent:chloroform:methanol:water = 50:10:1, v / v / v), so it is determined that post-treatment is possible.

[0405] The reaction mixture is rapidly cooled with methanol (30 mL), concentrated under reduced pressure (approximately 20 mL), and precipitated with water (200 mL). The crude reaction product is filtered through a sintered glass filter and thoroughly washed with water (3 × 300 mL). The crude material is dried in a drying box in the presence of KOH and P2O5 until it reaches a certain weight (recovered material: 10.1 g).

[0406] The crude reaction product is purified by chromatography, and the fraction containing the product is collected based on TLC analysis. This fraction is evaporated under reduced pressure until dry to obtain a white substance. This is dried in a drying box in the presence of KOH and P2O5 until it reaches a constant weight (TBDMS-αCD-butyl-βCD-TBDMS dimer, 3.6 g).

[0407] Step 2: Deprotection of the butyl-linked TBDMS-αCD-βCD dimer

[0408] Under an inert atmosphere, anhydrous TBDMS-αCD-butyl-βCD-TBDMS dimer (3.6 g, 0.98 mmol) was dissolved in THF (250 mL), and tetrabutylammonium fluoride (9.63 g, 36.85 mmol) was added all at once to the yellowish solution. After stirring at room temperature for 30 minutes, the reaction mixture turned dark green. The reaction mixture was stirred at room temperature overnight. TLC analysis (1,4-dioxane:25% NH3 (aqueous solution) = 10:7 (v / v)) revealed that the reaction was not complete, so the second volume of tetrabutylammonium fluoride (4 g, 13.3 mmol) was added to the container. The reaction mixture was gently heated until reflux was achieved and continued for 2 hours. At this stage, the reaction transformation was complete, as there were no starting materials detectable by TLC. The reaction mixture was cooled to room temperature, concentrated under reduced pressure (to approximately 10 mL), and methanol (200 mL) was added to obtain a white precipitate. The solid was filtered off and analyzed by TLC, then dried in a drying box in the presence of KOH and P2O5 until a constant weight was obtained (1.1 g). The mother liquor was concentrated under reduced pressure (to approximately 10 mL), purified by chromatography (eluent: 1,4-dioxane: 25% NH3 (aqueous solution) = 10:7 v / v), and the fraction containing the product was collected based on TLC analysis. This was evaporated under reduced pressure until dry to obtain a white substance. This was dried in a drying box in the presence of KOH and P2O5 until a constant weight was obtained (αCD-butyl-βCD dimer, 0.55 g).

[0409] Step 3: Hydroxypropylation of αCD-butyl-βCD heterodimer

[0410] Suspend αCD-butyl-βCD dimer (0.55 g, 0.25 mmol) in water (10 mL), and add sodium hydroxide (0.1 g, 2.5 mmol) to the reaction vessel. The mixture will become a slightly yellow solution. Cool the reaction mixture in a water bath (10°C) and add propylene oxide (0.5 mL, 0.415 g, 7.14 mmol) all at once. Flash the reaction vessel with argon, seal it, and stir at room temperature for 2 days. Concentrate the reaction mixture under reduced pressure until a viscous syrup is obtained, and precipitate it with acetone (50 mL). Filter the white solid through a sintered glass filter and wash thoroughly with acetone (3 × 15 mL). Dissolve this substance in water (50 mL), treat with ion exchange resin (to remove salts), clarify with activated carbon, filter by membrane, and dialyze against purified water for 1 day. The remaining amount is evaporated under reduced pressure until dry to obtain a white solid (0.63 g).

[0411] Detailed explanation of the synthesis of HP(αCD-triazole-βCD) heterodimer and HP(βCD-triazole-αCD) heterodimer.

[0412] Step 3a: Synthesis of αCD-triazole-βCD dimer

[0413] While stirring vigorously, suspend 2-O-monopropargyl-αCD and 2-O-mono(3-azidopropyl)-βCD in water (300 mL) (each at a concentration of approximately 8-12 mM). To completely dissolve the heterogeneous mixture, add approximately 300 mL of N,N-dimethylformamide to the suspension (adding DMF is a slightly exothermic process). Add copper bromide (2 g, 13.49 mmol) to the solution. Stir the suspension at room temperature for 1 hour. The reaction is monitored by TLC and is expected to be complete after approximately 1 hour (eluent: CH3CN:H2O:NH3 = 10:5:2). Filter the crude reaction product and concentrate the mother liquor under reduced pressure (60°C). Dilute the gel-like substance with water and add silica (15 g). Concentrate the heterogeneous mixture under reduced pressure until dry. This crude mixture is placed on the top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution). After drying, αCD-triazole-βCD dimer is obtained.

[0414] Step 3b: Synthesis of βCD-triazole-αCD dimer

[0415] While stirring vigorously, suspend 2-O-monopropargyl-βCD and 2-O-mono(3-azidopropyl)-αCD in water (300 mL) (each at a concentration of approximately 8-12 mM). To completely dissolve the heterogeneous mixture, add N,N-dimethylformamide (approximately 300 mL) to this suspension (addition of DMF is a slightly exothermic process). Add copper bromide (2 g, 13.49 mmol) to the solution. Stir the suspension at room temperature for 1 hour. The reaction is monitored by TLC and is expected to be complete after approximately 1 hour (eluent: CH3CN:H2O:NH3 = 10:5:2). Filter the crude reaction product and concentrate the mother liquor under reduced pressure (60°C). Dilute the gel-like substance with water and add silica (15 g). Concentrate the heterogeneous mixture under reduced pressure until dry. This crude mixture is placed on top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% v / v NH3 aqueous solution). After drying, βCD-triazole-αCD dimer is obtained.

[0416] Step 4a: HP(αCD-triazole-βCD) dimer

[0417] Suspend αCD-triazole-βCD dimer (1 g, 0.418 mmol), obtained by steps 1-3 above or by other methods, in water (50 mL). Add sodium hydroxide (DS3 = 0.32 g, 8 mmol; DS6 = 0.74 g, 18.5 mmol; DS7 = 0.87 g, 21.75 mmol) to the reaction vessel, and the mixture will become a slightly yellow solution. Cool the reaction mixture in a water bath (10°C) and add propylene oxide (DS3 = 0.49 mL, 0.42 g, 7.25 mmol; DS6 = 1.21 mL, 1.04 g, 17.9 mmol; DS7 = 1.46 mL, 1.7 g, 29.3 mmol) all at once. Flash the reaction vessel with argon, seal it, and stir at room temperature for 2 days. The solution is concentrated under reduced pressure until a viscous syrup is obtained, and this is precipitated with acetone (50 mL). The white solid is filtered through a sintered glass filter and thoroughly washed with acetone (3 × 15 mL). This substance is dissolved in water (50 mL), treated with ion exchange resin (to remove salts), clarified with activated carbon, filtered through membrane filtration, and dialyzed against purified water for 1 day. The residue is evaporated under reduced pressure until dry to obtain a white solid (0.7 g).

[0418] Step 4b: HP(βCD-triazole-αCD) dimer

[0419] Suspend βCD-triazole-αCD dimer (1 g, 0.418 mmol), obtained by steps 1-3 above or by other methods, in water (50 mL). Add sodium hydroxide (DS3=0.32 g, 8 mmol; DS6=0.74 g, 18.5 mmol; DS7=0.87 g, 21.75 mmol) to the reaction vessel, and the mixture will become a slightly yellow solution. Cool the reaction mixture in a water bath (10°C) and add propylene oxide (DS3=0.49 mL, 0.42 g, 7.25 mmol; DS6=1.21 mL, 1.04 g, 17.9 mmol; DS7=1.46 mL, 1.7 g, 29.3 mmol) all at once. Flash the reaction vessel with argon, seal it, and stir at room temperature for 2 days. The solution was concentrated under reduced pressure until a viscous syrup was obtained, and this was precipitated with acetone (50 mL). The white solid was filtered through a sintered glass filter and thoroughly washed with acetone (3 × 15 mL). This substance was dissolved in water (50 mL), treated with ion exchange resin (to remove salts), clarified with activated carbon, filtered through membrane filtration, and dialyzed against purified water for 1 day. The residue was evaporated under reduced pressure until dry to obtain a white solid (0.6 g).

[0420] Example 6. Synthesis of methyl-substituted CD dimers

[0421] Figure 7M shows the molecules that are synthesized.

[0422] This example describes the synthesis of a methyl-substituted CD dimer having a triazole-containing linker.

[0423] Methyl (βCD-triazole-βCD) dimer (synthesis example)

[0424] The preparation of the methylated β-CD dimer was achieved in a single step (see Figure 7K). The βCD-triazole-βCD dimer nucleus was prepared by the synthetic strategy described in Example 5 above.

[0425] synthesis

[0426] While vigorously stirring, βCD-triazole-βCD dimer nuclei (1.1 g, 0.46 mmol) were suspended in deionized H2O (100 mL), and sodium hydroxide (0.35 g, 8.8 mmol) was added. The resulting slightly yellow suspension was stirred for 30 minutes until completely dissolved. When the temperature of the yellowish clear solution stabilized at approximately 20°C, methyl iodide (0.5 mL, 1.14 g, 8.03 mmol) was added all at once while vigorously stirring (Note: Methyl iodide is immiscible with the reaction mixture, therefore, vigorous stirring was performed to efficiently convert it). The reaction mixture was stirred at room temperature for 24 hours, and then treated with an ion exchange resin (H in the solution). + Resin (6g) and OH - (6g) resin was added, stirred for 15 minutes, and filtered (the resin was washed with 3 × 15 mL of deionized water). The resulting filtrate (final pH = 7) was clarified with activated carbon (while stirring vigorously, activated carbon (0.2g) was added to the solution, stirred for 30 minutes, and filtered (the activated carbon pad was washed with 3 × 15 mL of deionized water)). The colorless solution was evaporated under reduced pressure (40°C) to obtain the title compound as a white powder (approximately 1g).

[0427] Characterization

[0428] The reaction process was monitored by TLC, and the resulting materials were characterized by MALDI-TOF (Figure 7L) and NMR analysis, as shown in Figures 7N, 7O, and 7P.

[0429] Example 7. Synthesis of sulfobutyl-substituted CD dimers

[0430] Figure 7S shows the molecules that are synthesized.

[0431] This example describes the synthesis of a sulfobutyl-substituted CD dimer having a triazole-containing linker.

[0432] The SB dimer was prepared in a single-step reaction (Figure 7Q).

[0433] Synthesis of SB(βCD-triazole-βCD) dimer low DS

[0434] While vigorously stirring, βCD-triazole-βCD dimer nuclei (1.2 g, 0.5 mmol) were suspended in deionized H2O (60 mL). Sodium hydroxide (0.39 g, 9.75 mmol) was added to the mixture, and the resulting solution was heated to 60°C. 1,4-butanesultone (0.88 mL, 1.17 g, 8.6 mmol) was added dropwise at 60°C, and the solution was heated at the same temperature for 3 hours. Next, to decompose the unreacted 1,4-butanesultone, the reaction mixture was further heated to 90°C for 1 hour. The reaction mixture was cooled and treated with an ion exchange resin. A cation exchange resin (H) was added to the solution. + Resin, 2g) and anion exchange resin (OH - Add 2g of resin, stir for 15 minutes, and filter (wash the resin with 3 x 15 mL of deionized water). The resulting filtrate (final pH=7) was clarified with activated carbon (while stirring vigorously, add 0.3g of activated carbon to the solution, stir for 30 minutes, and filter (wash the activated carbon pad with 3 x 15 mL of deionized water)).

[0435] The colorless solution was evaporated under reduced pressure (40°C) to obtain a white powder (1.47 g).

[0436] Characterization

[0437] The reaction was monitored by TLC analysis, and the resulting substances were characterized by MALDI-TOF (Figure 7R) and NMR analysis, as shown in Figures 7T-7Z.

[0438] Synthesis of SB(βCD-triazole-βCD) dimer high DS

[0439] While vigorously stirring, (βCD-triazole-βCD) dimer nuclei (1.2 g, 0.5 mmol) were suspended in deionized H2O (60 mL). Sodium hydroxide (1.22 g, 30.5 mmol) was added to the mixture, and the resulting solution was heated to 60°C. 1,4-butanesultone (2.8 mL, 3.72 g, 27.35 mmol) was added dropwise at 60°C, and the solution was heated at the same temperature for 3 hours. Then, to decompose the remaining 1,4-butanesultone, the reaction mixture was further heated at 90°C for 1 hour. The reaction mixture was cooled and treated with an ion exchange resin. A cation exchange resin (H) was added to the solution. + Resin, 4g) and anion exchange resin (OH - Add 4g of resin and stir for 15 minutes, then filter (wash the resin with 3 x 15 mL of deionized water). The resulting filtrate (final pH = 7) was clarified with activated carbon (while stirring vigorously, add 0.5g of activated carbon to the solution, stir for 30 minutes, and filter (wash the activated carbon pad with 3 x 15 mL of deionized water)). The colorless solution was evaporated under reduced pressure (40°C) to obtain a white powder (1.51g).

[0440] Characterization

[0441] The obtained materials were characterized by MALDI-TOF (Figure 7W) and NMR analysis, as shown in Figures 7X to 7Z.

[0442] Example 8. Synthesis of quaternary ammonium-substituted CD dimers

[0443] Figure 7AC shows the molecules that are synthesized.

[0444] This example describes the synthesis of a quaternary ammonium-substituted CD dimer having a triazole-containing linker.

[0445] Quaternary ammonium (βCD-triazole-βCD) dimer (synthesis example)

[0446] The preparation of the QA dimer was achieved in a single step (see Figure 7AA). The βCD-triazole-βCD dimer nucleus was prepared by the synthetic strategy described in Example 5 above.

[0447] QA(β-CD-triazole-β-CD) dimer (Synthesis example)

[0448] While stirring vigorously, the β-CD-triazole-β-CD dimer nucleus (1.2 g, 0.5 mmol) was suspended in deionized H2O (100 mL), and sodium hydroxide (0.39 g, 9.8 mmol) was added. The resulting slightly yellow suspension was stirred for 30 minutes until completely dissolved. When the temperature of the yellowish transparent solution was stable at 5 °C to 10 °C, glycidyltrimethylammonium chloride (1.17 mL, 1.32 g, 8.7 mmol) was added at once while stirring vigorously. The reaction mixture was stirred at room temperature for 24 hours, and then, when the temperature of the solution was stable at 5 °C to 10 °C, a second portion of glycidyltrimethylammonium chloride (0.4 mL, 0.45 g, 3 mmol) was added. The reaction mixture was heated at 50 °C for 3 hours, then cooled and treated with an ion exchange resin (H + resin (6 g) and OH - (6 g) resin were added, stirred for 15 minutes, and filtered (the resin was washed with 3 × 15 mL of deionized water)). The resulting filtrate (final pH = 7) was clarified with activated carbon (while stirring vigorously, activated carbon (0.2 g) was added to the solution, stirred for 30 minutes, and filtered off (the activated carbon pad was washed with 3 × 15 mL of deionized water)). The colorless solution was evaporated under reduced pressure (40 °C) to obtain the title compound as a white powder (about 800 mg).

[0449] Characterization

[0450] The resulting substance was characterized by MALDI-TOF (Figure 7AB) and NMR analysis as shown in Figures 7AD to 7AF.

[0451] In the case of QA-BCD derivatives, the typical Gaussian distribution with a regular pattern observed in randomly substituted derivatives is lost during MALDI analysis, while an irregular pattern of fragmentation is detected. Identifying / assigning these irregular peaks is difficult because a simple pattern of fragmentation cannot be predicted. The most likely cause of the irregular pattern observed in the MALDI spectrum is the instability of the trimethylammonium moiety under experimental conditions. Specifically, the detachment product is the result of partial cleavage of trimethylammonium, while the demethylation product is the result of progressive cleavage of the methyl group from the cation side chain. Since peaks with no informational value are generated during laser detachment, it is reasonable to conclude that the MALDI conditions are not suitable for identifying the DS of QA-βCD derivatives. However, the DS of QA-βCD derivatives can be identified by NMR (Figure 7AD), and it was estimated to be approximately 2.1.

[0452] Example 9. Synthesis of succinyl-substituted CD dimers

[0453] Figure 7AI shows the synthesized molecule. The succinyl-substituted dimer (Succ dimer) was prepared in a single step (Figure 7AG).

[0454] Synthesis of succinyl-substituted cyclodextrin dimers

[0455] Under an inert atmosphere, βCD-triazole-βCD dimer nuclei (1.2 g, 0.5 mmol) were suspended in pyridine (23 mL) with vigorous stirring. The suspension was heated at 40°C for 1 hour to increase the solubility of the βCD-triazole-βCD dimer nuclei, but complete dissolution was not achieved. A second dose of pyridine (23 mL) was added to the suspension, but dilution did not further improve the solubility of the βCD-triazole-βCD dimer nuclei. Succinic anhydride (0.1 g, 1 mmol) was added at room temperature, and the reaction mixture was stirred for 24 hours. The crude reaction product was concentrated under reduced pressure, dissolved in water (50 mL) (a clear solution was not obtained), and treated with an ion exchange resin (H2O2 was added to the solution). + Resin (2g) and OH- (2g) resin was added, stirred for 15 minutes, and filtered (the resin was washed with 3 × 15 mL of deionized water). The resulting filtrate (final pH = 7) was clarified with activated carbon (while stirring vigorously, activated carbon (0.5g) was added to the solution, stirred for 30 minutes, and filtered (the activated carbon pad was washed with 3 × 15 mL of deionized water)). The colorless solution was evaporated under reduced pressure (40°C) to obtain the title compound as a white powder (approximately 900 mg).

[0456] Characterization

[0457] The obtained materials were characterized by MALDI-TOF (Figure 7AH) and NMR analysis, as shown in Figures 7AJ to 7AL.

[0458] Similar to the case of the QA dimer, MALDI analysis proved unsuitable for identifying the DS, and the DS was identified by NMR (Figure 7AD), estimated to be approximately 2.1.

[0459] Detailed explanation of the synthesis of HPβCD-triazole-βCD (random substitution) asymmetric dimers.

[0460] The preparation of the HPβCD-triazole-βCD DS3 (random substitution) asymmetric dimer is achieved through multiple synthetic steps, as shown in Figure 20A.

[0461] Step 1: Preparation of Azidrinker

[0462] Dissolve 1,3-dibromopropane (10 mL, 20.18 g, 0.1 mol) in 40 mL of DMSO while stirring vigorously. Prepare a DMSO (240 mL) solution of sodium azide (6.7 g, 0.1 mol) and add it dropwise to the 1,3-dihalopropane solution (over 2 hours). Stir the solution overnight at room temperature. Next, extract the crude reaction product with n-hexane (3 × 100 mL), extract the collected organic phase with water (3 × 50 mL), and carefully evaporate the resulting organic phase under reduced pressure (strictly at 40°C and 400 mbar; otherwise, the target compound may be removed by distillation). Purify the oily residue by chromatography (isocratic elution with n-hexane-siRNA = 98:2 as the eluent). Based on TLC analysis, appropriate fractions are collected and concentrated under reduced pressure to obtain the target compound as a viscous oily substance (which can be stored in a light-shielded refrigerated container under an inert atmosphere). The compound is visualized by immersing the TLC plate in a 10% triphenylphosphine solution in dichloromethane for approximately 15 seconds, drying the TLC plate at below 60°C, immersing the TLC in a 2% ninhydrin ethanol solution for approximately 15 seconds, and finally drying the TLC plate at below 60°C. The target compound appears as purple spots on the TLC plate.

[0463] Step 2.1: Preparation of 2-O-monopropargyl-βCD

[0464] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred under N2 at room temperature for 12-24 hours until clear. Then, propargyl bromide (1.97 mL, 17.62 mmol) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% NH3 aqueous solution) shows spots corresponding to monopropargyl and nonpropargyl βCD, respectively. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture was placed on the top of a silica column (25 × 6 cm) and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After freeze-drying, 2-O-monopropargyl-βCD was obtained as a solid. 2-O-monopropargyl-βCD was analyzed by MALDI and NMR.

[0465] Step 2.2: Random (2-hydroxypropyl)-2-O-monopropargyl-βCD

[0466] The 2-O-monopropargyl-βCD (4.9, 4.2 mmol) obtained in Step 2.1 was suspended in water (500 mL), and sodium hydroxide (DS3 = 3.2 g, 80 mmol) was added to the reaction vessel, resulting in a slightly yellow solution. The reaction mixture was cooled in a water bath (10°C), and propylene oxide (DS3 = 4.9 mL, 0.4.2 g, 72.5 mmol) was added all at once. The reaction vessel was flushed with argon, sealed, and stirred at room temperature for 2 days. The solution was concentrated under reduced pressure until a viscous syrup was obtained, and this was precipitated with acetone (50 mL). The white solid was filtered through a sintered glass filter and thoroughly washed with acetone (3 × 15 mL). This substance was dissolved in water (50 mL), treated with ion exchange resin (to remove salts), clarified with activated carbon, membrane filtered, and dialyzed against purified water for 1 day. The residue was evaporated under reduced pressure until dry. Random (2-hydroxypropyl)-2-O-monopropargyl-βCD was separated as a white solid (4.2 g) and analyzed by NMR (Figures 22A-22D). The degree of substitution was calculated by NMR spectroscopy (Figure 22B).

[0467] Step 2.3: Synthesis of 2-O-mono(3-azidopropyl)-βCD

[0468] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred at room temperature under N2 until clear (12-24 hours). Then, 3-azido-1-bromopropane (3 mL) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)) shows spots corresponding to 2-O-mono(3-azidopropyl)-βCD and βCD. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture was placed on top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After drying, 2-O-mono(3-azidopropyl)-βCD was obtained as a white solid.

[0469] Step 3: Synthesis of HPβCD'-triazole-βCD (random substitution) asymmetric dimer

[0470] While vigorously stirring, suspend random (2-hydroxypropyl)-2-O-monopropargyl-βCD and 2-O-mono(3-azidopropyl)-βCD in water (300 mL) (each at a concentration of approximately 8-12 mM). To completely dissolve the heterogeneous mixture, add approximately 300 mL of N,N-dimethylformamide (DMF) to the suspension (addition of DMF is a slightly exothermic process). Add copper bromide (2 g, 13.49 mmol) to the solution. Stir the suspension at room temperature for 1 hour. Monitor the reaction by TLC and expect it to be complete after approximately 1 hour (eluent: CH3CN:H2O:25% NH3 (aqueous solution) = 10:5:2). Filter the crude reaction product and concentrate the mother liquor under reduced pressure (60°C). Dilute the gel-like substance with water and add silica (15 g). Concentrate the heterogeneous mixture under reduced pressure until dry. This crude mixture was placed on the top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After drying, randomly substituted HPβCD-triazole-βCD asymmetric dimer DS3 was obtained. The dimer preparation was characterized by NMR.

[0471] Detailed explanation of the synthesis of the C6HPβCD-triazole-βCD DS3 asymmetric dimer.

[0472] The preparation of the C6HPβCD-triazole-βCD DS3 asymmetric dimer can be achieved through multiple synthetic steps, as shown in Figures 20B-20D.

[0473] Step 1A: Preparation of Azidrinker

[0474] Dissolve 1,3-dibromopropane (10 mL, 20.18 g, 0.1 mol) in 40 mL of DMSO while stirring vigorously. Prepare a DMSO (240 mL) solution of sodium azide (6.7 g, 0.1 mol) and add it dropwise to the 1,3-dihalopropane solution (over 2 hours). Stir the solution overnight at room temperature. Next, extract the crude reaction product with n-hexane (3 × 100 mL), extract the collected organic phase with water (3 × 50 mL), and carefully evaporate the resulting organic phase under reduced pressure (strictly at 40°C and 400 mbar; otherwise, the target compound may be removed by distillation). Purify the oily residue by chromatography (isocratic elution with n-hexane-siRNA = 98:2 as the eluent). Based on TLC analysis, appropriate fractions are collected and concentrated under reduced pressure to obtain the target compound as a viscous oily substance (which can be stored in a light-shielded refrigerated container under an inert atmosphere). The compound is visualized by immersing the TLC plate in a 10% triphenylphosphine solution in dichloromethane for approximately 15 seconds, drying the TLC plate at below 60°C, immersing the TLC in a 2% ninhydrin ethanol solution for approximately 15 seconds, and finally drying the TLC plate at below 60°C. The target compound appears as purple spots on the TLC plate.

[0475] Step 1B: Preparation of a protected 2-hydroxypropylating agent (1-bromo-2-benzyloxypropane)

[0476] 2-benzyloxy-1-propanol

[0477] Acid-catalyzed alcohol decomposition was carried out using a round-bottom flask, reflux condenser, thermometer, and stepwise drop funnel. Benzyl alcohol containing the catalyst (sulfuric acid) was heated to the reaction temperature, and propylene oxide was added as quickly as the reflux rate allowed. After the addition, heating was continued until the boiling point reached a constant temperature, indicating that the olefin oxide had been consumed. The catalyst was neutralized with sodium hydroxide, and the product was isolated by fractional distillation. Specifically, 63.8 g (1.1 mol) of propylene oxide was added to 600 g (5.55 mol) of benzyl alcohol containing 1 g of sulfuric acid over 4 hours, while maintaining the liquid temperature at 120°C to 125°C. After further heating over 2 hours, the temperature was stabilized at 120°C. Approximately 77 g of 2-benzyloxy-1-propanol was obtained from the mixture.

[0478] 1-Bromo-2-benzyloxy-propane

[0479] 2-benzyloxy-1-propanol (16.6 g, 0.1 mol) is solubilized in ACN (100 mL), and slowly added to the ACN suspension of P2O5 (21.3 g, 0.15 mol) and KBr (17.85 g, 0.15 mol) under an inert atmosphere with vigorous stirring (added over 20 minutes). The reaction mixture is stirred at room temperature for 3 hours, then concentrated under reduced pressure (approximately 10 mL). The suspension is solubilized in water under cooling (0°C to 5°C) and neutralized with sodium carbonate. The resulting mixture is extracted using DCM (3 × 100 mL), and the organic phase is combined and concentrated under reduced pressure to obtain a viscous yellowish oil. The residue is purified by silica gel chromatography (20% siRNA / n-hexane) to obtain 1-bromo-2-benzyloxy-propane (17.7 g, 85%) as a colorless oil. 1H NMR (500MHz, CDCl3)δ:1.33 (3H, d, J=6.7Hz, CH3), 3.39 (1H, dd, J=4.9, 10.4Hz, CH2Br), 3.46 (1H, dd, J=4.9, 10.4Hz, CH2Br), 3.73-3.76 (1H, m, CH), 4.59 (2H, s, PhCH2), 7.27-7.38 (5H, m, phenyl). 13C NMR (126MHz, CDCl3)δ:19.2, 36.8, 71.2, 74.3, 127.9, 128.6, 138.4.

[0480] Step 2.1: Preparation of 2-O-monopropargyl-βCD

[0481] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.43 mmol) is added. The resulting suspension is stirred under N2 at room temperature for 12-24 hours until clear. Then, propargyl bromide (1.97 mL, 17.62 mmol) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% NH3 aqueous solution) shows spots corresponding to monopropargyl and nonpropargyl βCD, respectively. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture was placed on the top of a silica column (25 × 6 cm) and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After freeze-drying, 2-O-monopropargyl-βCD was obtained as a solid. 2-O-monopropargyl-βCD was analyzed by MALDI and NMR.

[0482] Step 2.2: Per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD

[0483] 2-O-monopropargyl-βCD (10 g, 8.5 mmol) was suspended in dry pyridine (200 mL) under N2 and stirred at room temperature for 30 minutes until a clear solution was formed. Then, tert-butyldimethylsilyl chloride (10.8 g, 71.4 mmol) was added all at once, and the resulting suspension was stirred at room temperature for 6 hours. TLC (15:2:1 ethyl-96% v / v EtOH-H2O) yielded the title product (R f The formation of (=0.65) was observed, showing both low-polarity and high-polarity substances corresponding to insufficiently silylated and excessively silylated species, respectively. Part of TBDMSCl (2.7 g, 17.9 mmol) was added every 6 hours until the aforementioned spots completely disappeared (Figure 23A). The solution was then poured into a mixture of 5% HCl aqueous solution (3 L) and ice, stirred until the ice melted, and extracted with CH2Cl2 (2 × 4 L). The combined organic phase was washed with H2O (2 × 4 L), dried (MgSO4), and concentrated. Trace amounts of pyridine were removed by co-evaporation with toluene (2 × 3 L). The obtained substance was mixed with silica gel (30 g), suspended in CH2Cl2, and the solvent was removed under vacuum. First, an aqueous solution of NH3 with CH2Cl2-CH3CN-96% v / v and EtOH-30% v / v in a 40:40:20:4 ratio was used as the eluent, and R f Chromatography was performed using TLC (15:2:1 siRNA-96%v / v EtOH-H2O) spots at 0.87 and 0.80 until the compound was eluted (2L). Subsequently, per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD was dried under high vacuum at 100°C for 6 hours using 40:40:20:4 CH2Cl2-CH3CN-96%v / v EtOH-H2O (1.5L) as the solvent to obtain a white amorphous powder (12.6g, 6.4 mmol, 75%). The material was decomposed at 232°C-236°C. [α] 25 D +96 (c1.0, CH2Cl2). R f= 0.64 (15:2:1 EtOAc - 96% v / v EtOH - H₂O). IR (ATR) (Figure 23F): 3313, 2953, 2930, 2887, 2857, 1253, 1155, 1083, 1038, 833, 777, 735 cm -1 (Trotta, F.; Martina, K.; Robaldo, B.; Barge, A.; Cravotto, G. J. Incl. Phenom. Macrocyclic Chem. 2007, 57, 3 - 7) (KBr) 3420, 3325, 1473, 1254, 1086, 1040, 835 cm -1 ; 1 ¹H NMR (Figures 23C - 23D) (500 MHz, CDCl₃) δ 5.34 (bs, OH), 5.05 (d, 1H, 3 J 1, 2= 3.2 Hz, H - 1 I ), 4.89 - 4.88 (m, 6H, H - 1 II-VII ), 4.50 (dd, 1H, 2 ​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​), 79.6 (C≡), 75.2 (≡CH), 74.0-72.5 (C2 I-VII , 3 I-VII , 5 I-VII ), 62.3-61.6 (C-6 I-VII ), 59.9 (CH2C≡), 26.0[SiC (CH3)3], 18.5-18.3 [SiC (CH3)3], -4.9- (-5.1) (SiCH3). MALDI-TOF (Figure 23B): [M+Na] + C 87 H 170 O 35 Calculated value for Si7Na: 1995.0, measured value: 1995.0, C 87 H 170 O 35 Analytical calculation values ​​for Si7: C, 52.97, H, 8.69. Measured values: C, 52.93, H, 8.71. [ka] [ka] [ka] [ka]

[0484] Step 2.3: per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD

[0485] Dissolve per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD (12.6 g, 6.4 mmol) in THF (500 mL). Cool the solution to 10°C in an ice bath, and add KOH (129 g, 1.98 mol) in small amounts while stirring vigorously. The resulting white suspension is initially slightly viscous, but then becomes easier to stir. Methyltriphenylphosphonium bromide (5.04 g, 14 mmol) was added to the reaction mixture, and the white suspension was stirred for 3 hours. Benzyl bromide (46.1 mL, 66.4 g, 0.39 mmol) was slowly and carefully added to the heterogeneous mixture while maintaining the temperature below 25°C (over 2 hours). After stirring for 1 hour, the reaction mixture became pearly white with a milky viscosity. This reaction was stirred overnight at room temperature. The progress of the reaction was monitored by TLC (n-hexane:siRNA = 9:1), the reaction mixture was cooled to 10°C, and the second additions of KOH (12.96 g, 0.23 mol) and BnBr (4.6 mL, 6.64 g, 0.04 mol) were added. After 2 hours, the third additions of KOH (6.5 g, 0.12 mol) and BnBr (2.3 mL, 3.3 g, 0.02 mmol) were added, and the reaction mixture was stirred for a further 3 hours. The heterogeneous mixture was filtered through a sintered glass filter (porosity 4), and the solid was thoroughly washed with THF (3 × 200 mL). The filtrate was concentrated in a rotor vaporizer (approximately 50 mL) and poured into MeOH (500 mL) with vigorous stirring. The resulting yellowish gel-like substance was separated by decantation. The solid was thoroughly washed with H2O (5 × 600 mL) and MeOH:H2O=1:9 (3 × 400 mL), and finally dried in a vacuum drying box in the presence of P2O5 and KOH as desiccants until it reached a constant weight. Per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD was isolated as a white powder (42 g, 80%).

[0486] Step 2.4: per-2,3-O-benzyl-2-O-monopropargyl-βCD

[0487] Under an inert atmosphere, per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD (42 g) was solubilized in THF (800 mL), and tetrabutylammonium fluoride trihydrate (19.75 g, 0.062 mol) was added in small amounts. The yellowish solution was stirred overnight at room temperature. Desilylation was followed by TLC (CHCl3:MeOH=9:1), which was completed overnight. The crude reaction product was concentrated using a rotary vaporizer, methanol (600 mL) was added, and the solution was concentrated again using a rotary vaporizer. The azeotropic distillation procedure was repeated three times (3 × 600 mL methanol), and finally the crude product was concentrated until dry. The remaining yellowish substance was suspended in water (1 L), filtered through a sintered glass filter (porosity 4), and thoroughly washed with water (5 × 300 mL) and a mixture of MeOH:H2O=1:9 (3 × 300 mL) until a white, odorless solid was obtained. The white solid was dried in a vacuum drying box in the presence of P2O5 and KOH as desiccants until it reached a certain weight. Per-2,3-O-benzyl-2-O-monopropargyl-βCD was isolated as a white solid (21 g, 8.96 mmol).

[0488] Step 2.5: Tris(6-O-(2-O-benzyloxypropyl))-per-2,3-O-benzyl-2-O-monopropargyl-βCD

[0489] Per-2,3-O-benzyl-2-O-monopropargyl-βCD (21 g, 8.96 mmol) was dissolved in THF (300 mL). The solution was cooled to 10°C in an ice bath, and KOH (2.5 g, 44.8 mmol) was added in small amounts while vigorously stirring. The resulting white suspension was initially slightly viscous, but then became easier to stir. Methyltriphenylphosphonium bromide (0.5 g, 1.4 mmol) was added to the reaction mixture, and the white suspension was stirred for 3 hours. While keeping the temperature below 25°C, 1-bromo-2-benzyloxypropane (10.3 g, 44.8 mmol) was slowly added to the heterogeneous mixture. After stirring for 1 hour, the reaction mixture became pearly white with a milky viscosity. This reaction was stirred overnight at room temperature. The progress of the reaction was monitored by TLC (n-hexane:siRNA = 9:1). The heterogeneous mixture was filtered through a sintered glass filter (porosity 4), and the solid was thoroughly washed with THF (3 × 50 mL). The filtrate was concentrated in a rotor vaporizer (approximately 30 mL) and poured into MeOH (200 mL) with vigorous stirring. The resulting yellowish gel-like substance was separated by decantation. The solid was thoroughly washed with H2O (5 × 100 mL) and MeOH:H2O=1:9 (3 × 100 mL). Tris(6-O-(2-O-benzyloxypropyl))-per-2,3-O-benzyl-2-O-monopropargyl-βCD was purified by chromatography using isocratic elution with silica gel (n-hexane:Â=9:1). The fractions were combined based on TLC analysis and concentrated to dryness under reduced pressure. Tris(6-O-(2-O-benzyloxypropyl))-per-2,3-O-benzyl-2-O-monopropargyl-βCD was isolated as a white powder (15 g, 5.4 mmol, 60%).

[0490] Step 2.6: Tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD

[0491] Tris(6-O-(2-O-benzyloxypropyl))-per-2,3-O-benzyl-2-O-monopropargyl-βCD (15 g, 5.4 mmol) was solubilized in methanol (500 mL). The reaction mixture was heated at 40°C, and Pd / C (3.1 g) was added while vigorously stirring. Hydrazine carbonate (120 mL) was added dropwise to the container (over 1.5 hours). The mixture was heated gently under reflux for 3 hours, and the progress of the reaction was monitored by TLC (1,4-dioxane:25% NH3 (aqueous solution):1-propanol = 10:7:3). The reaction mixture was cooled to room temperature and filtered through a sintered glass filter (porosity 3). The Pd / C pad was thoroughly washed with MeOH (3 × 300 mL), H2O (3 × 300 mL), and MeOH:H2O = 50:50 (3 × 300 mL). The filtrate was evaporated to dryness using a rotor vaporizer (60°C). The residual solid was solubilized in water (120 mL), treated with an ion exchange resin, and purified with activated carbon. The resulting solution was then filtered through a Celite pad and finally evaporated to dryness. The solid was dried in a vacuum drying box in the presence of P2O5 and KOH as desiccants until it reached a constant weight. Tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD was isolated as a white powder (6 g, 4.45 mmol, 82%).

[0492] Step 2.7: Synthesis of 2-O-mono(3-azidopropyl)-βCD

[0493] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred at room temperature under N2 until clear (12-24 hours). Then, 3-azido-1-bromopropane (3 mL) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)) shows spots corresponding to 2-O-mono(3-azidopropyl)-βCD and βCD. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture was placed on top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After drying, 2-O-mono(3-azidopropyl)-βCD was obtained as a white solid.

[0494] Step 3: Synthesis of C6HPβCD-triazole-βCD DS3 asymmetric dimer

[0495] While stirring vigorously, suspend tris-6-O-(2-O-hydroxypropyl)-2-O-monopropargyl-βCD and 2-O-mono(3-azidopropyl)-βCD in water (300 mL) (each at a concentration of approximately 8-12 mM). To completely dissolve the heterogeneous mixture, add N,N-dimethylformamide (DMF) (approximately 300 mL) to this suspension (addition of DMF is a slightly exothermic process). Add copper bromide (2 g, 13.49 mmol) to the solution. Stir the suspension at room temperature for 1 hour. The reaction is monitored by TLC and is expected to be complete after approximately 1 hour (eluent: CH3CN:H2O:25% NH3 (aqueous solution) = 10:5:2). Filter the crude reaction product and concentrate the mother liquor under reduced pressure (60°C). Dilute the gel-like substance with water and add silica (15 g). The heterogeneous mixture was concentrated under reduced pressure until dry. This crude mixture was placed on the top of a silica column and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After drying, the C6HPβCD-triazole-βCD asymmetric dimer DS3 was obtained.

[0496] Detailed explanation of the synthesis of the C6HPβCD-triazole-βCD DS7 asymmetric dimer.

[0497] The preparation of the C6HPβCD-triazole-βCD DS7 asymmetric dimer was achieved through multiple synthetic steps, as shown in Figures 21A-21D.

[0498] Step 1A: Preparation of Azidrinker

[0499] Dissolve 1,3-dibromopropane (10 mL, 20.18 g, 0.1 mol) in 40 mL of DMSO while stirring vigorously. Prepare a DMSO (240 mL) solution of sodium azide (6.7 g, 0.1 mol) and add it dropwise to the 1,3-dihalopropane solution (over 2 hours). Stir the solution overnight at room temperature. Next, extract the crude reaction product with n-hexane (3 × 100 mL), extract the collected organic phase with water (3 × 50 mL), and carefully evaporate the resulting organic phase under reduced pressure (strictly at 40°C and 400 mbar; otherwise, the target compound may be removed by distillation). Purify the oily residue by chromatography (isocratic elution with n-hexane-siRNA = 98:2 as the eluent). Based on TLC analysis, appropriate fractions are collected and concentrated under reduced pressure to obtain the target compound as a viscous oily substance (which can be stored in a light-shielded refrigerated container under an inert atmosphere). The compound is visualized by immersing the TLC plate in a 10% triphenylphosphine solution in dichloromethane for approximately 15 seconds, drying the TLC plate at below 60°C, immersing the TLC in a 2% ninhydrin ethanol solution for approximately 15 seconds, and finally drying the TLC plate at below 60°C. The target compound appears as purple spots on the TLC plate.

[0500] Step 1B: Preparation of a protected 2-hydroxypropylating agent (1-bromo-2-benzyloxypropane)

[0501] 2-benzyloxy-1-propanol

[0502] Acid-catalyzed alcohol decomposition was carried out using a round-bottom flask, reflux condenser, thermometer, and stepwise drop funnel. Benzyl alcohol containing the catalyst (sulfuric acid) was heated to the reaction temperature, and propylene oxide was added as quickly as the reflux rate allowed. After the addition, heating was continued until the boiling point reached a constant temperature, indicating that the olefin oxide had been consumed. The catalyst was neutralized with sodium hydroxide, and the product was isolated by fractional distillation. Specifically, 63.8 g (1.1 mol) of propylene oxide was added to 600 g (5.55 mol) of benzyl alcohol containing 1 g of sulfuric acid over 4 hours, while maintaining the liquid temperature at 120°C to 125°C. After further heating over 2 hours, the temperature was stabilized at 120°C. From the mixture, 77 g of 2-benzyloxy-1-propanol was obtained.

[0503] 1-Bromo-2-benzyloxy-propane

[0504] 2-benzyloxy-1-propanol (16.6 g, 0.1 mol) was solubilized in ACN (100 mL) and slowly added to an ACN suspension of P2O5 (21.3 g, 0.15 mol) and KBr (17.85 g, 0.15 mol) under an inert atmosphere with vigorous stirring (added over 20 minutes). The reaction mixture was stirred at room temperature for 3 hours and then concentrated under reduced pressure (approximately 10 mL). The suspension was solubilized in water under cooling (0°C to 5°C) and neutralized with sodium carbonate. The resulting mixture was extracted using DCM (3 × 100 mL), and the organic phase was combined and concentrated under reduced pressure to obtain a viscous yellowish oil. The residue was purified by silica gel chromatography (20% ethyl acetate / n-hexane) to obtain 1-bromo-2-benzyloxy-propane (17.7 g, 85%) as a colorless oil. 1H NMR (500MHz, CDCl3) δ:1.33 (3H, d, J=6.7Hz, CH3), 3.39 (1H, dd, J=4.9, 10.4Hz, CH2Br), 3.46 (1H, dd, J=4.9, 10.4Hz, CH2Br), 3.73-3.76 (1H, m, CH), 4.59 (2H, s, PhCH2), 7.27-7.38 (5H, m, phenyl). 13C NMR (126MHz, CDCl3) δ:19.2, 36.8, 71.2, 74.3, 127.9, 128.6, 138.4.

[0505] Step 2.1: Preparation of 2-O-monopropargyl-βCD

[0506] To a solution of βCD (20 g, 17.62 mmol) in anhydrous DMSO (300 mL), lithium hydride (212 mg, 26.432 mmol) is added. The resulting suspension is stirred under N2 at room temperature for 12-24 hours until clear. Then, propargyl bromide (1.97 mL, 17.62 mmol) and a catalytic amount of lithium iodide (approximately 20 mg) are added, and the mixture is stirred at 55°C for 5 hours in the dark. Characterization of the product using TLC (10:5:2 CH3CN-H2O-25% NH3 aqueous solution) shows spots corresponding to monopropargyl and nonpropargyl βCD, respectively. The solution is poured into acetone (3.2 L), the precipitate is filtered, and the mixture is thoroughly washed with acetone. The resulting solid is transferred to a round-bottom flask and dissolved in the minimum volume of water. Silica gel (40 g) is added, and the solvent is removed under vacuum until a powdery residue is obtained. This crude mixture was placed on the top of a silica column (25 × 6 cm) and subjected to chromatography (10:5:2 CH3CN-H2O-25% NH3 (aqueous solution)). After freeze-drying, 2-O-monopropargyl-βCD was obtained as a solid. 2-O-monopropargyl-βCD was analyzed by MALDI and NMR.

[0507] Step 2.2: Per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD

[0508] 2-O-monopropargyl-βCD (10 g, 8.5 mmol) was suspended in dry pyridine (200 mL) under N2 and stirred at room temperature for 30 minutes until a clear solution was formed. Then, tert-butyldimethylsilyl chloride (10.8 g, 71.4 mmol) was added all at once, and the resulting suspension was stirred at room temperature for 6 hours. TLC (15:2:1 ethyl-96% v / v EtOH-H2O) yielded the title product (R f The formation of (=0.65) was observed, showing both low-polarity and high-polarity substances corresponding to insufficiently silylated and excessively silylated species, respectively. Part of TBDMSCl (2.7 g, 17.9 mmol) was added every 6 hours until the aforementioned spots completely disappeared (Figure 23A). The solution was then poured into a mixture of 5% HCl aqueous solution (3 L) and ice, stirred until the ice melted, and extracted with CH2Cl2 (2 × 4 L). The combined organic phase was washed with H2O (2 × 4 L), dried (MgSO4), and concentrated. Trace amounts of pyridine were removed by co-evaporation with toluene (2 × 3 L). The obtained substance was mixed with silica gel (30 g), suspended in CH2Cl2, and the solvent was removed under vacuum. First, a 40:40:20:4 CH2Cl2-CH3CN-96% v / v EtOH-30% v / v NH3 aqueous solution was used as the eluent, R f Chromatography was performed using TLC (15:2:1 siRNA-96%v / v EtOH-H2O) spots at 0.87 and 0.80 until the compound was eluted (2L). Subsequently, per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD was dried under high vacuum at 100°C for 6 hours using 40:40:20:4 CH2Cl2-CH3CN-96%v / v EtOH-H2O (1.5L) as the solvent to obtain a white amorphous powder (12.6g, 6.4 mmol, 75%). The material was decomposed at 232°C-236°C. [α] 25 D +96 (c1.0, CH2Cl2). R f= 0.64 (EtOAc - 96% v / v EtOH - H2O in a ratio of 15:2:1). IR (ATR) (Figure 23F): 3313, 2953, 2930, 2887, 2857, 1253, 1155, 1083, 1038, 833, 777, 735 cm -1 (Trotta, F.; Martina, K.; Robaldo, B.; Barge, A.; Cravotto, G. J. Incl. Phenom. Macrocyclic Chem. 2007, 57, 3 - 7) (KBr) 3420, 3325, 1473, 1254, 1086, 1040, 835 cm -1 ; 1 1H NMR (Figures 23C - 23D) (500 MHz, CDCl3) δ 5.34 (bs, OH), 5.05 (d, 1H, 3 J 1, 2= 3.2 Hz, H - 1 I ), 4.89 - 4.88 (m, 6H, H - 1 II-VII ), 4.50 (dd, 1H, 2 J = 16.7 Hz, 4 J = 2.3 Hz, CHO), 4.41 (dd, 1H, 2 J = 16.7 Hz, 4 J = 2.3 Hz, CHO), 4.11 - 3.82 (m, 14H, H - 3 I-VII S, 6a I-VII ), 3.74 - 3.49 (m, 28H, H - 2 I-VII , 4 I-VII , 5 I-VII , 6b I-VII ), 2.40 (t, 1H, 4 J = 2.3 Hz, ≡CH), 0.88 - 0.86 [m, 63H, SiC(CH3)3], 0.04 - 0.​​​​​​​​​), 79.6 (C≡), 75.2 (≡CH), 74.0 - 72.5 (C2 I-VII , 3 I-VII , 5 I-VII ), 62.3 - 61.6 (C - 6 I-VII ), 59.9 (CH2C≡), 26.0 [SiC(CH3)3], 18.5 - 18.3 [SiC(CH3)3], -4.9 - (-5.1) (SiCH3). MALDI - TOF (Figure 23B): [M + Na] + C 87 H 170 O 35 Calculated value for Si7Na: 1995.0, Measured value: 1995.0, C 87 H 170 O 35 Analytical calculated value for Si7: C, 52.97, H, 8.69. Measured value: C, 52.93, H, 8.71.

[0509] Step 2.3: per - 6 - O - tert - butyldimethylsilyl - per - 2,3 - O - benzyl - 2 - O - monopropargyl - βCD

[0510] Dissolve per-6-O-tert-butyldimethylsilyl-2-O-monopropargyl-βCD (12.6 g, 6.4 mmol) in THF (500 mL). Cool the solution to 10°C in an ice bath and add KOH (129 g, 1.98 mol) in small amounts while stirring vigorously. The resulting white suspension is initially slightly viscous but then becomes easy to stir. Add methyltriphenylphosphonium bromide (5.04 g, 14 mmol) to the reaction mixture and stir the white suspension for 3 hours. Slowly and carefully add benzyl bromide (46.1 mL, 66.4 g, 0.39 mmol) to the heterogeneous mixture while keeping the temperature below 25°C (over 2 hours). After stirring for 1 hour, the reaction mixture becomes pearly white with a milky viscosity. Stir this reaction mixture overnight at room temperature. The reaction was monitored by TLC (n-hexane:HCl = 9:1), the reaction mixture was cooled to 10°C, and the second additions of KOH (12.96 g, 0.23 mol) and BnBr (4.6 mL, 6.64 g, 0.04 mol) were added. After 2 hours, the third additions of KOH (6.5 g, 0.12 mol) and BnBr (2.3 mL, 3.3 g, 0.02 mmol) were added, and the reaction mixture was stirred for a further 3 hours. The heterogeneous mixture was filtered through a sintered glass filter (porosity 4), and the solid was thoroughly washed with THF (3 × 200 mL). The filtrate was concentrated in a rotor vaporizer (approximately 50 mL) and poured into MeOH (500 mL) while vigorously stirring. The resulting yellowish gel-like substance was separated by decantation. The solid was thoroughly washed with H2O (5 × 600 mL) and MeOH:H2O=1:9 (3 × 400 mL), and finally dried in a vacuum drying box in the presence of P2O5 and KOH as desiccants until it reached a constant weight. Per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD was isolated as a white powder (42 g, 80%).

[0511] Step 2.4: per-2,3-O-benzyl-2-O-monopropargyl-βCD

[0512] Under an inert atmosphere, per-6-O-tert-butyldimethylsilyl-per-2,3-O-benzyl-2-O-monopropargyl-βCD (42 g) is solubilized in THF (800 mL), and tetrabutylammonium fluoride trihydrate (19.75 g, 0.062 mol) is added in small amounts. The yellowish solution is stirred at room temperature overnight. Desilylation is followed by TLC (CHCl3:MeOH=9:1) and completed overnight. The crude reaction mixture is concentrated using a rotary vaporizer, methanol is added (600 mL), and the solution is concentrated again using a rotary vaporizer. The azeotropic distillation procedure is repeated three times (3 × 600 mL methanol), and finally the crude product is concentrated until dry. The remaining yellowish substance is suspended in water (1 L) and filtered through a sintered glass filter (porosity 4). The mixture is then thoroughly washed with water (5 × 300 mL) and a mixture of MeOH:H2O=1:9 (3 × 300 mL) until a white, odorless solid is obtained. The white solid is dried in a vacuum drying box in the presence of P2O5 and KOH as desiccants until it reaches a certain weight. Per-2,3-O-benzyl-2-O-monopropargyl-βCD is isolated as a white solid (21 g, 8.96 mmol).

[0513] Step 2.5: per-6-O-(2-O-benzyloxypropyl)-per-2,3-O-benzyl-2-O-monopropargyl-βCD

[0514] Dissolve per-2,3-O-benzyl-2-O-monopropargyl-βCD (21 g, 8.96 mmol) in THF (300 mL). Cool the solution to 10°C in an ice bath and add KOH (25 g, 448 mmol) in small amounts while stirring vigorously. The resulting white suspension will be sligh...

Claims

1. The following general formula structure A-X: CD-[A-B-A']-CD' (Structure AX) A cyclodextrin (CD) dimer having, in the formula, CD is a βCD having structure A-Xa: 【Chemistry 1】 In the formula, CD' is a βCD having structure A-Xb: 【Chemistry 2】 During the ceremony, L1, L2, L3, L1', L2', and L3' are each identical or different, and each is independently selected from the group consisting of bond, -O-, -NH-, -NR4-, and -S-. At least one of L1, L2, L3, L1', L2', and / or L3' is not O or is a bond. [A-B-A'] is defined as a linking group that together comprises a linker length of 2 to 20 atoms, and CD and CD' are linked via a secondary plane by at least one of the aforementioned linking groups. A and A' are independently selected from the group consisting of a bond, -O-, -NH-, -NR4-, -S-, a heteroatom, an alkylene, or an alkylene in which at least one carbon atom is substituted by a heteroatom selected from the list consisting of O, N, S, Si, and P. B is selected from the group consisting of bonds, -O-, -NH-, -NR4-, -S-, heteroatoms, alkylenes, alkylenes in which at least one carbon atom is substituted by a heteroatom selected from the list consisting of O, N, S, Si, or P, saturated or unsaturated cycloalkylenes, saturated or unsaturated heterocycloalkylenes, arylenes, and heteroarylenes. At least one of A, B, and A' of each linking group is not a bond, Each linking group A, B, or A' is connected to at least one L1 or L2, and the corresponding R1 or R2 is omitted. Each linking group A, B, or A' is connected to at least one L1' or L2', and the corresponding R1' or R2' is omitted. R1, R2, R3, R4, R1', R2', and R3' are either identical or different, and each is independent of the others: Hydrogen, methyl, 2-hydroxypropyl, 3-hydroxypropyl, -CH ２ CH(OH)CH ２ N(CH ３ ) ３ ＋ Group (I) consisting of sulfobutyl, succinyl, carboxymethyl, maltosyl, glucosyl, acetyl, amino, ammonium, azide, 1,2-ethylenediamine, carboxy, carbamoyl, carboxamide, cyano, fluoro, hydroxy, sulfuric acid, sulfonamide, trimethylammonium propyl, ureid, SH, F, Cl, Br, and I; or Contains 1 to 6 carbon atoms, including alkyl, hydroxyalkyl, sulfoalkyl, trialkylammoniumalkyl, hydroxytrialkylammoniumalkyl, carboxyalkyl, alkenyl, alkynyl, alkoxy, alkoxyalkyl, alkoxyalkoxyalkyl, alkylcarbonyl, alkylcarbonyloxyalkyl, alkoxycarbonyloxy, hydroxyalkoxy, hydroxyalkoxyalkyl, hydroxycarbonylalkyl, alkylamino, dialkylamino, trialkylammonium, aminoalkyl, alkylaminoalkyl, dialkylaminoalkyl, aminoalkoxy, hydroxyalkylamino, hydroxyalkylaminoalkyl, aminocarbonyl Group (II) consisting of oxyalkyl, alkoxyamino, alkoxycarbonylamino, alkylaminocarbonyl, alkylaminocarbonylalkyl, alkylsulfonylamide, aminosulfonyl, alkylaminosulfonyl, dialkylaminosulfonyl, alkylsulfanyl, thioalkyl, alkylsulfonyl, alkylsulfonylalkyl, alkylsulfonamide, cyanoalkyl, alkylureido, cycloalkyl, heterocyclyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkylalkyl, cycloalkoxy, heterocycloalkoxy, heterocycloamino, heterocyclylalkyl, heterocyclyloxy, and hydroxycycloalkyl; Selected from, and The CD dimer wherein at least one of R1, R2, R3, R1', R2', and R3' that is not linked to a linking group is not hydrogen and contains one of the substituents selected from group (I) or group (II).

2. The following general formula structure B-X or structure B-X': CD-[A-B-A']-CD' (Structure B-X) CD'-[A-B-A']-CD (Structure B-X') A CD dimer having, in the formula, The CD is an αCD having the following structure B-Xa, 【Transformation 3】 In the formula, CD' is a βCD having the following structure B-Xb, 【Chemistry 4】 During the ceremony, L1, L2, L3, L1', L2', and L3' are each identical or different, and each is independently selected from the group consisting of bond, -O-, -NH-, -NR4-, and -S-. [A-B-A'] is defined as a linking group that together comprises a linker length of 2 to 20 atoms, and CD and CD' are linked via a secondary plane by at least one of the aforementioned linking groups. A and A' are independently selected from the group consisting of a bond, -O-, -NH-, -NR4-, -S-, a heteroatom, an alkylene, and an alkylene in which at least one carbon atom is substituted by a heteroatom selected from the list consisting of O, N, S, Si, and P. B is selected from the group consisting of a bond, -O-, -NH-, -NR4-, -S-, heteroatom, alkylene, saturated or unsaturated cycloalkylene, saturated or unsaturated heterocycloalkylene, arylene, and heteroarylene. At least one of A, B, and A' of each linking group is not a bond, Each linking group A, B, or A' is connected to at least one L1 or L2, and the corresponding R1 or R2 is omitted. Each linking group A, B, or A' is connected to at least one L1' or L2', and the corresponding R1' or R2' is omitted. R1, R2, R3, R4, R1', R2', and R3' are either identical or different, and each is independent of the others: Hydrogen, methyl, 2-hydroxypropyl, 3-hydroxypropyl, -CH ２ CH(OH)CH ２ N(CH ３ ), ３ ＋ , a group (I) consisting of sulfobutyl, succinyl, carboxymethyl, maltosyl, glucosyl, acetyl, amino, ammonium, azide, 1,2-ethylenediamine, carboxy, carbamoyl, carboxamide, cyano, fluoro, hydroxy, sulfuric acid, sulfonamide, trimethylammoniumpropyl, ureido, SH, F, Cl, Br, and I; or Contains 1 to 6 carbon atoms, including alkyl, hydroxyalkyl, sulfoalkyl, trialkylammoniumalkyl, hydroxytrialkylammoniumalkyl, carboxyalkyl, alkenyl, alkynyl, alkoxy, alkoxyalkyl, alkoxyalkoxyalkyl, alkylcarbonyl, alkylcarbonyloxyalkyl, alkoxycarbonyloxy, hydroxyalkoxy, hydroxyalkoxyalkyl, hydroxycarbonylalkyl, alkylamino, dialkylamino, trialkylammonium, aminoalkyl, alkylaminoalkyl, dialkylaminoalkyl, aminoalkoxy, hydroxyalkylamino, hydroxyalkylaminoalkyl, aminocarbonyl Group (II) consisting of oxyalkyl, alkoxyamino, alkoxycarbonylamino, alkylaminocarbonyl, alkylaminocarbonylalkyl, alkylsulfonylamide, aminosulfonyl, alkylaminosulfonyl, dialkylaminosulfonyl, alkylsulfanyl, thioalkyl, alkylsulfonyl, alkylsulfonylalkyl, alkylsulfonamide, cyanoalkyl, alkylureido, cycloalkyl, heterocyclyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkylalkyl, cycloalkoxy, heterocycloalkoxy, heterocycloamino, heterocyclylalkyl, heterocyclyloxy, and hydroxycycloalkyl; The CD dimer selected from the above.

3. (a) R1, R2, R3, R1', R2', and R3' are either identical or different, and each is independently selected from group (I) or group (II); and at least two R3 and R3' are not hydrogen; (b) R1, R2, R3, R1', R2', and R3' are either identical or different, and each is independently selected from group (I) or group (II); and at least two and no more than four R3 and R3' are not hydrogen; (c) R1, R1', R2, and R2' are each hydrogen; R3 and R3' are each the same or different, each independently selected from group (I) or group (II); (d) R1, R1', R2, and R2' are each hydrogen; R3 and R3' are each the same or different; and each is independently selected from group (I) or group (II); and at least two R3 and R3' are not hydrogen; (e) R1, R1', R2, and R2' are each hydrogen; R3 and R3' are each the same or different; and each is independently selected from group (I) or group (II); and at least two and four or fewer R3 and R3' are not hydrogen; (f) R3 and R3' are each hydrogen; R1, R1', R2, and R2' are each the same or different, each independently selected from group (I) or group (II); or (g) R3 and R3' are identical groups selected from group (I); and R1, R1', R2, and R2' may be identical or different, and each is independently selected from group (I) or group (II); A CD dimer according to any one of claims 1 to 2.

4. (a) The DS at position C2 of CD (corresponding to L1-R1) is not equal to the degree of substitution (DS) at position C2 of CD' (corresponding to L1'-R1'). (b) The DS at position C3 of CD (corresponding to L2-R2) is not equal to the DS at position C3 of CD' (corresponding to L2'-R2'). (c) The DS at position C6 of CD (corresponding to L3-R3) is not equal to the DS at position C6 of CD' (corresponding to L3'-R3'). (d) At least one L1-R1, L2-R2, or L3-R3 pair is different from the respective L1'-R1', L2'-R2', and L3'-R3' pairs. (e) at least one L1'-R1', L2'-R2', or L3'-R3' pair is different from the respective L1-R1, L2-R2, and L3-R3 pairs, or (f) Any combination of (a) to (e) A CD dimer according to any one of claims 1 to 3, comprising at least one of the above.

5. A CD dimer according to any one of claims 1 to 4, wherein at least one of R1, R2, R3, R1', R2', or R3' is not hydrogen, and the corresponding L1, L2, L3, L1', L2', or L3' is not O or contains a bond.

6. In the linking group [A-B-A'], B is structure Y: 【Transformation 5】 A and A', or structure Y', as shown: 【Transformation 6】 It contains a triazole having connectivity to A'-A as shown in, and A and A' are each independently alkylenes having a length of 1 to 8 carbon atoms; or A is methyl and A' is propyl, the CD dimer according to any one of claims 1 to 5.

7. The CD dimer according to any one of claims 1 to 6, wherein at least one of R1, R2, R3, R1', R2', and / or R3' is methyl, 2-hydroxypropyl, sulfobutyl, succinyl, maltosyl, carboxymethyl, trimethylammoniumpropyl, or 2-hydroxytrimethylammoniumpropyl.

8. The following general formula structure B-X or structure B-X': CD-[A-B-A']-CD' (Structure B-X) CD'-[A-B-A']-CD (Structure B-X') A CD dimer having, in the formula, The CD is an αCD having the following structure B-Xa, 【Transformation 7】 In the formula, CD' is a βCD having the following structure B-Xb, 【Transformation 8】 During the ceremony, L1, L2, L3, L1', L2', and L3' are each identical or different, and each is independently selected from the group consisting of bond, -O-, -NH-, -NR4-, or -S-. R1, R2, R3, R4, R1', R2', and R3' are each hydrogen. [A-B-A'] together is defined as a linking group, and is linked to the secondary surfaces of CD and CD'. A and A' are alkylenes having a length of 1 to 8 carbon atoms, respectively; or A is methyl and A' is propyl. B is structure Y: 【Chemistry 9】 A and A', or structure Y', as shown: 【Chemistry 10】 It contains a triazole having connectivity from A' to A as shown in, A or A' of each linking group is connected to at least one L1 or L2, and the corresponding R1 or R2 is omitted, and The CD dimer, wherein A or A' of each linking group is connected to at least one L1' or L2', and the corresponding R1' or R2' is omitted.

9. A CD dimer composition comprising a mixture of two or more CD dimers according to any one of claims 1 to 8, wherein the CD dimer composition optionally substantially contains other CD dimers.

10. A pharmaceutical composition comprising a CD dimer according to any one of claims 1 to 8, or a CD dimer composition according to claim 9 and a pharmaceutically acceptable carrier.

11. (a) comprising the CD dimer or the CD dimer composition and the pharmaceutically acceptable carrier, or essentially comprising them, and / or (b) The pharmaceutical composition according to claim 10, wherein the CD dimer or the CD dimer composition is the sole active ingredient in the pharmaceutical composition.

12. The pharmaceutical composition according to claim 10, further comprising at least one hydrophobic drug comprising estrogen, progesterone, and / or testosterone, and optionally comprising the CD dimer or the CD composition in an amount effective for solubilizing the hydrophobic drug.

13. A pharmaceutical composition according to any one of claims 10 to 12 for reducing the amount of 7-ketocholesterol (7KC) and / or cholesterol in a subject requiring treatment.

14. Atherosclerosis / coronary artery disease, arteriosclerosis, coronary artery atherosclerosis due to calcified coronary artery lesions, heart failure (all stages), Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, Huntington's disease, vascular dementia, multiple sclerosis, Smith-Lemle-Oppitz syndrome, pediatric neuronal ceroid lipofuscinosis, lysosomal acid lipase deficiency, cerebral tendon xanthomatous dystrophy, X-linked adrenoleukodystrophy, sickle cell disease, Niemann-Pick disease type A, Niemann-Pick disease type B, The pharmaceutical composition according to claim 13, which prevents, treats, or alleviates symptoms of one or more of the following: Niemann-Pick disease type C, Gaucher disease, Stargardt disease, age-related macular degeneration (atrophic type), idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, cystic fibrosis, liver injury, liver failure, non-alcoholic steatohepatitis, non-alcoholic fatty liver disease, irritable bowel syndrome, Crohn's disease, ulcerative colitis, and / or hypercholesterolemia, and optionally, the treatment is administered in combination with another treatment.

15. A method for improving the solubility of a hydrophobic drug, comprising estrogen, progesterone, and / or testosterone, the method comprising mixing the hydrophobic drug with a CD dimer according to any one of claims 1 to 8, a CD dimer composition according to claim 9, or a pharmaceutical composition according to claim 10.

16. Use of a CD dimer according to any one of claims 1 to 8, a CD dimer composition according to claim 9, or a pharmaceutical composition according to claim 10 in the manufacture of a pharmaceutical for reducing the amount of 7KC and / or cholesterol in a subject requiring treatment.

17. A method for producing a CD dimer according to any one of claims 1 to 8, (a) A primary-protected CD molecule is reacted with a dialkylating agent to produce a primary-protected CD dimer linked via a secondary surface, and the primary-protected CD dimer is optionally purified. (b) Deprotecting the primary surface protected CD dimer to produce a deprotected CD dimer, and optionally purifying the deprotected CD dimer. (c) The method comprising optionally functionalizing the deprotected CD dimer with the R1, R2, R3, R1', R2', and / or R3' groups to produce the CD dimer, and optionally purifying the CD dimer.

18. (i) The CD protected on the primary surface comprises one or more protecting groups selected from the group consisting of trityl, benzoyl, tert-butyldimethylsilyl (TBDMS), tert-butyldiphenylsilyl (TBDPS), and triisopropylsilyl (TIPS), or comprises heptakis(6-O-tert-butyldimethylsilyl)cyclodextrin; (ii) The dialkylating agent comprises a dihaloalkane or 1,4-dibromobutane; (iii) Step (a) is carried out under anhydrous conditions and / or using sodium hydride as the base; (iv) The purification in step (a) comprises normal-phase or reverse-phase chromatography using isocratic elution and / or crystallization; (v) Step (b) is carried out using tetrabutylammonium fluoride in tetrahydrofuran (THF); (vi) The purification in step (b) includes normal-phase or reverse-phase chromatography using isocratic elution and / or crystallization; or (vii) Step (c) comprises reacting the deprotected CD dimer with a hydroxypropylating agent comprising propylene oxide, a methylating agent comprising methyl iodide, a succinylating agent comprising succinic anhydride, a sulfobutylating agent comprising 1,4-butanesultone, and / or a quaternary ammonium reagent comprising glycidyltrimethylammonium chloride; (viiii) Step (c) is carried out in an aqueous solvent with a base comprising sodium hydroxide or lithium hydroxide; and / or (ix) The purification in step (c) comprises one or more of the following: ion exchange resin treatment, activated carbon clarification, and dialysis; or Any combination of (x)(i) to (ix); The method according to claim 17, including the method described in claim 17.

19. A method for producing a CD dimer according to any one of claims 1 to 8, comprising (a) reacting 2-O-(n-azidoalkyl)-CD or 3-O-(n-azidoalkyl)-CD or a mixture thereof with 2-O-(n-alkyne)-CD or 3-O-(n-alkyne)-CD or a mixture thereof to form a CD-triazole-CD dimer, and optionally comprising (b) purifying the CD-triazole-CD dimer.

20. (i) Step (a) is carried out using a copper(I) catalyst, a silver(I) catalyst, a ruthenium catalyst, a copper(I) bromide, or copper bromide tris(triphenylphosphine) [(PPh3)3CuBr]; (ii) Step (a) is carried out in an aqueous solution or in a solution containing water and 50% dimethylformamide; (iii) The purification in step (b) comprises silica gel chromatography and / or crystallization; or (iv) The method further comprises, prior to step (a), reacting an n-azido-1-bromoalkane with a base comprising cyclodextrin and lithium hydride, sodium hydride, and n-butyllithium, and optionally with a catalytic amount of lithium iodide in DMSO to produce the 2-O-(n-azidoalkyl)-CD; (v) The method further comprises purifying the 2-O-(n-azidoalkyl)-CD by silica gel chromatography; (vi) The method further comprises, prior to step (a), reacting the n-bromo-1-alkyne with a base comprising cyclodextrin and lithium hydride, sodium hydride, and n-butyllithium, and optionally with a catalytic amount of lithium iodide in DMSO to produce the 2-O-(n-alkyne)-CD; (vii) The method further comprises purifying the 2-O-(n-alkyne)-CD by silica gel chromatography; (viiii) The method further comprises, after step (b), reacting the CD-triazole-CD dimer with a base comprising lithium hydroxide or sodium hydroxide and one or more of the following: a hydroxypropylating agent comprising propylene oxide, a methylating agent comprising methyl iodide, a succinylating agent comprising succinic anhydride, a sulfobutylating agent comprising 1,4-butanesultone, and / or a quaternary ammonium reagent comprising glycidyltrimethylammonium chloride; (ix) Purifying the CD-triazole-CD dimer by one or more of the following: ion exchange resin treatment, activated carbon clarification, membrane filtration, and dialysis; or Any combination of (x)(i) to (ix); The method according to claim 19, including the method described in claim 19.

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