TREATMENT OF SUBSTANCE ADDICTION

CROSS-REFERENCE TO RELATED APPLICATION

[01] This application claims priority to U.S. Application No. 13/946,201, filed July 19, 2013, the disclosure of each of which is hereby incorporated by reference in its entirety.

BACKGROUND

[02] Combustion of organic materials such as tobacco is known to produce tar and other potentially harmful materials. Inhalers have been used as one non-combustion type of device for delivering nicotine for inhalation. Some types of inhalers use a highly pressurized chamber for delivering a powdered medium for inhalation. See, e.g., U.S. Patent 7,461,649 to Gamard et al. Other varieties of inhalers employ a tube in which the active component is volatized into vapor form for inhalation. See, e.g., U.S. Patent 6,769,436 to Horian. Current inhalers of this type generally use nicotine, a toxic and addictive substance, as the sole active component.

[03] The art has recognized a need for non-nicotine or reduced-nicotine alternatives to tobacco and tobacco replacement products. See D.K. Hatsukami et al., "Nicotine reduction revisited: science and future directions, ^' " Tobacco Control 19: el-elO (2010). There remains a need for non-nicotine or reduced-nicotine alternatives that may assist smokers in quitting traditional cigarette smoking. It would be particularly desirable to develop a product that is useful in treating nicotine addiction and/or other substance addictions, while avoiding or minimizing the individual's exposure to nicotine.

SUMMARY

[04] Embodiments of the present invention are directed to inhalers and similar devices adapted for delivering anatabine in a volatilized form. In one aspect, a volatilized form of anatabine is delivered for treating a substance addiction such as nicotine addiction, cocaine addiction, marijuana addiction, heroin addiction, or alcohol addiction.

BRIEF DESCRIPTION OF THE DRAWINGS

[05] A more complete understanding of the present invention and certain advantages thereof may be acquired by referring to the following detailed description in consideration with the accompanying drawings, in which:

[06] FIG. 1 is a schematic illustration of an inhaler in accordance with one embodiment.

[07] FIG. 2 is a sectional view of the drum of the inhaler shown in FIG. 1 used to hold the alkaloid composition.

[08] FIG. 3 is a schematic illustration of an inhaler in accordance with another embodiment.

[09] FIG. 4 shows that administration of single doses of anatabine dose-dependently decreased nicotine self-administration.

[10] FIG. 5 shows the nicotine dose-effect curve in monkeys under the same conditions as in FIG. 4.

[11] FIG. 6 shows the effects of anatabine on nicotine self-administration by rats.

DETAILED DESCRIPTION

[12] Tobacco is among the most chemically complex substances known, with tobacco and tobacco smoke containing more than 8,000 compounds. In addition to nicotine, tobacco contains the minor alkaloids nornicotine, anabasine, and anatabine. While nicotine is regarded as the principal addictive component in tobacco, a variety of other factors also are believed to contribute to tobacco addiction. For example, tobacco smoke has been reported to have a monoamine oxidase (MAO) inhibitory effect. MAO is an enzyme involved in the breakdown of dopamine, a pleasure-enhancing neurotransmitter. See J.S. Fowler et al., "Inhibition of Monoamine Oxidase B in the Brain of Smokers," Nature (Lond), 379(6567):733 736 (1996); J. Stephenson, "Clues Found to Tobacco Addiction," Journal of the American Medical Association, 275(16): 1217-1218 (1996). See also Williams et al. U.S. Patent 6,350,479.

[13] Aspects of the present invention are directed to inhalers and similar devices that deliver anatabine in a volatilized form for treating substance addiction while avoiding or reducing the subject's exposure to nicotine. In some examples, anatabine is the sole alkaloid present in the composition, e.g., anatabine comprises 100 wt. % of the total alkaloid weight. In other examples, up to about 75 wt. % of one or more other alkaloids, such as nicotine, nornicotine, and/or anabasine, may be present in addition to anatabine. For example, anatabine and nicotine may be combined in a weight ratio (anatabine-to-nicotine) of about 50: 1 to about 1 :3, or from about 25: 1 to about 1 :2, from about 10: 1 to about 3:2, or from about 5: 1 to about 1 : 1. In some examples, anatabine is the only active component present. In other examples, anatabine may be combined with other active component(s).

[14] Unless otherwise clear from context, all percentages referred to herein are expressed as percent by weight based on the total weight of the composition.

[15] Anatabine may be prepared synthetically, such as via a benzophenoneimine pathway as described in U.S. Patent 8,207,346 Bl, the disclosure of which is incorporated herein by reference in its entirety. Anatabine may be present in the form of a racemic mixture or as isolated enantiomer, e.g., R-(+)-anatabine or S-(-)-anatabine, and/or as one or more pharmaceutically acceptable (or food grade) salts of anatabine. Unless otherwise clear from context, "anatabine" as used herein refers to racemic mixtures of anatabine, enantiomers of anatabine, salt and non-salt forms of anatabine, as well as salt and non-salt forms of anatabine enantiomers. Non- limiting examples of possible salts are described in P. H. Stahl et al., Handbook of Pharmaceutical Salts: Properties, Selection and Use, Weinheim/Zurich:Wiley-VCH/VFICA, 2002, including salts of l-hydroxy-2-naphthoic acid, 2,2-dichloroacetic acid, 2-hydroxyethanesulfonic acid, 2-oxoglutaric acid, 4-acetamidobenzoic acid, 4-aminosalicylic acid, acetic acid, adipic acid, ascorbic acid (L), aspartic acid (L), benzenesulfonic acid, benzoic acid, camphoric acid (+), camphor- 10-sulfonic acid (+), capric acid (decanoic acid), caproic acid (hexanoic acid), caprylic acid (octanoic acid), carbonic acid, cinnamic acid, citric acid, cyclamic acid, dodecylsulfuric acid, ethane- 1 ,2-disulfonic acid, ethanesulfonic acid, formic acid, fumaric acid, galactaric acid, gentisic acid, glucoheptonic acid (D), gluconic acid (D), glucuronic acid (D), glutamic acid, glutaric acid, glycerophosphoric acid, glycolic acid, hippuric acid, hydrobromic acid, hydrochloric acid, isobutyric acid, lactic acid (DL), lactobionic acid, lauric acid, maleic acid, malic acid (- L), malonic acid, mandelic acid (DL), methanesulfonic acid, naphthalene- 1,5- disulfonic acid, naphthalene-2-sulfonic acid, nicotinic acid, nitric acid, oleic acid, oxalic acid, palmitic acid, pamoic acid, phosphoric acid, proprionic acid, pyroglutamic acid (- L), salicylic acid, sebacic acid, stearic acid, succinic acid, sulfuric acid, tartaric acid (+ L), thiocyanic acid, toluenesulfonic acid (p), and undecylenic acid.

[16] As an alternative to synthetic preparation, anatabine may be obtained by extraction from tobacco or other plants, such as members of the Solanaceae family, such as datura, mandrake, belladonna, capsicum, potato, tomato, nicotiana, eggplant, and petunia. A tobacco extract may be prepared from cured tobacco stems, lamina, or both. Flue (bright) varieties of tobacco are often used, i.e., Virginia flue. Other tobacco varieties may be used, such as Burley, dark-fired, and/or other commercial tobacco varieties. Two or more tobacco varieties may be combined to form a blend. In the extraction process, cured tobacco material is extracted with a solvent, typically water, ethanol, steam, or carbon dioxide. The resulting solution contains the soluble components of the tobacco, including alkaloids such as anatabine. Anatabine may be purified using known techniques such as liquid chromatography.

[17] The amount of anatabine present in the composition may vary depending on factors such as the type of device and/or whether other active components, such as nicotine and/or other alkaloids, are present. By way of example, the amount of anatabine may range from about 0.1 to about 25 mg, from about 0.5 to about 20 mg, or from about 1 to about 10 mg, per total gram of the composition.

[18] With respect to liquid compositions used for volatile delivery of anatabine as described herein, anatabine typically is present in an amount ranging from about 1 mg per lOmL to about 40 mg per lOmL of solution, often from about 4 mg per lOmL to about 30 mg per lOmL of solution, and sometimes from about 5 mg per lOmL to about 25 mg per lOmL of solution or from about 6 mg per lOmL to about 20 mg per lOmL of solution.

[19] As described in U.S. Patent 8,241,680 Bl to Williams et al, compositions containing anatabine were found to be efficacious for the temporary reduction of tobacco cravings, even without the presence of nicotine. Anatabine and other minor alkaloids also have been reported to bind to nicotinic receptors. See "Receptors for Nicotine in the Central Nervous System: 1 Radioligand Binding Studies," Group Research & Development Centre, British-American Tobacco Co. Ltd. (1984).

[20] In addition to anatabine, the composition may contain up to about 75 wt.% of one or more other alkaloids, such as nicotine, nornicotine, and/or anabasine, based on the total alkaloid weight. Such alkaloids may be extracted from tobacco or other plant materials and purified using known techniques, and/or prepared synthetically using known synthesis methods. Anatabine and additional alkaloid(s) may be combined in a weight ratio (anatabine -to-total other alkaloids) of about 50: 1 to about 1 :3, or from about 25: 1 to about 1 :2, from about 10: 1 to about 3 :2, or from about 5 : 1 to about 1 : 1.

[21] The composition may be provided in a solid, powdered form, or in liquid form, depending on such factors as the structure and operation of the particular device used for volatile delivery. Micronized dry powder used in inhalers typically is produced with an original particle range of about 1-10 microns. An individual dose may include, for example, from about 5 mg to about 20 mg of powder. The active agent(s) may be combined with one or more excipient carriers, non-limiting examples of which include lactose, trehalose, and mannitol. If desired, one or more flavorants may be added to the composition, non-limiting examples of which include peppermint, menthol, wintergreen, spearmint, propolis, eucalyptus, cinnamon, natural or artificial tobacco flavors, or the like. The total amount of flavorants and/or other additives typically ranges from about 0.5 to about 15 wt.%, often from about 1 to about 10 wt.%), based on the total weight of the composition.

[22] A liquid composition containing both anatabine and nicotine, for example, may contain a solvent together with anatabine in an amount of about 1 to about 40 mg per 10 mL of solution, often from about 3 to about 36 mg per 10 mL of solution, and nicotine in an amount of about 1 to about 40 mg per 10 mL of solution, often from about 9 to about 36 mg per 10 mL of solution. Suitable solvents include, but are not limited, to low-molecular-weight grades of polyethylene glycol (PEG), such as PEG 400 which has very low toxicity.

[23] The device used for volatile delivery may be of various types of configurations. By way of example and without limitation, volatile delivery of anatabine may be carried out by way of atomization, vaporization, use of a compressed gas, or other mechanical or electronic means (e.g., an electronic inhaler) capable of delivering anatabine in gaseous form or in a form otherwise suitable for inhalation.

[24] In some examples, solid forms of anatabine (such as a tablet, powder, or the like) may be dissolved in a suitable solvent or dispersed in a liquid vehicle. The anatabine- containing liquid may be filled into an inhaler or receptacle for a delivery device capable of administering anatabine in a volatilized form as described herein.

[25] In general, inhalers may be of a single-use or disposable type, or may be refillable with alkaloid compositions and/or cartridges containing alkaloid compositions to facilitate reuse. One example of an inhaler is shown in FIG. 1. The inhaler 30 of this embodiment has a high pressure chamber 32 coupled to an equalization chamber 34. The high pressure chamber 32 contains a compressed mixture 52 of helium and oxygen. The chamber 32 has a resealable, refilling opening 31. The high pressure chamber 32 includes a housing 36 defining a third chamber 38. The housing 36 includes an opening 40 on a top portion thereof and a gas passage 42 on a side. The third chamber 38 communicates with both the high pressure chamber 32 and the equalization chamber 34. A consistent volume of gas is produced with the help of a diaphragm plate 56.

[26] The equalization chamber 34 includes a housing 58 having a gasket 46 disposed therein. The gasket 46 includes a gas passage 48 on a side thereof for allowing gas disposed in the third chamber 38 to communicate with the second chamber 34. A piston 44 is slidably mounted within the gasket 46 and within the housing 36. The piston 44 includes a communication opening 50. The piston 44 is pushed downwards with a spring 60 located inside chamber 38 to allow gas communication between chambers 32 and 34. When the canister 32 is separated from the inhaler, the spring 60 is pushing the piston 44 sealing the canister by closing the opening 42. When the canister is inserted in the inhaler, the tip of the piston 44 will rest on the diaphragm 56, and pushing the piston 44 up inside chamber 38 just so that high-pressure gas passage 42 is communicating with the communication opening 50. The communication opening 50 is designed to selectively allow gas 52 stored in high pressure chamber 32 to communicate with gas stored in the equalization chamber 34. A pressure plate 56 is also disposed within housing 58. One side of pressure plate 56 is coupled to piston 44.

[27] Through the use of high pressure chamber 32 and equalization chamber 34, inhaler 30 produces a desired gas pressure without requiring an external pump. When the pressure inside the equalization chamber 34 is too low to allow inhaler 30 to be used, high-pressure gas 52 from the high pressure chamber 32 fills into equalization chamber 34. A spring 60 and pressure plate 56 are designed to facilitate this operation. The piston 44 has a communication opening 50 which selectively allows high pressure chamber 32 to communicate with equalization chamber 34 through gas passages 42 and 48.

[28] The gas 52 applies pressure against a small area defined by the top of piston 44. The net force applied by the high-pressure side of high pressure chamber 32 on the piston 44 works with the biasing force of spring 60 and against the force applied by gas 54 on pressure plate 56. The spring constant of spring 60 and the surface area of pressure plate 56 are chosen so that when equalization chamber 34 has received sufficient pressure to utilize inhaler 30, the force applied by the gas on pressure plate 56 will exceed that of the force produced by the gas 52 on piston 44 on the high- pressure side of the device and the force of the spring 60. At such a time, the force applied by the gas will cause piston 44 to move upward within housings 58 and 36. As piston 44 moves upwardly, communication opening 50 will move away from gas passage 48, effectively stopping any additional high-pressure gas 52 from entering equalization chamber 34. [29] Once the gas is dispensed, the pressure exerted by the gas on pressure plate 56 is lower and the high-pressure gas 52 along with the spring 60 will force piston 44 downwardly thereby repeating the cycle described above until equalization chamber 34 once again has a desired pressure of gas therein.

[30] One alternative to this delivery system employs mechanical actuation by the user. As the high-pressure chamber 32 is depressed, the piston 44 allows the gas to escape from the high-pressure gas passage 42 and be stored into a secondary chamber. The amount of gas released into equilibrium chamber 34 is then defined by the volume of this secondary chamber. The gas is released from this chamber into the equilibration chamber 34 when the high-pressure cylinder 32 is returned to its original position.

[31] An activating trigger 94 is disposed on a drum section 64. An inhalation port door 98 coupled to a mouth piece 99 of a spacer 96 closes to inhibit a user from inhaling the gas disposed within spacer 96. The spacer 96 may also have a scented receptacle 110 near to where a patient's nose would be. The air in the spacer 96 is purged through a pressure port 100.

[32] As the user inhales the dosage, a vacuum begins to form in spacer 96. At a certain pressure, vacuum/pressure valve 104 opens, allowing ambient air into the spacer 96. By opening vacuum/pressure valve 104, the user may continue a steady deep inhalation of ambient air following inhalation of the dosage.

[33] Upon actuation of the inhaler 30, the gas is injected into a gas passage 62 of a drum section 64. The drum section 64 includes a housing 65 that contains a rotating drum which includes a plurality of tubes 68 that are substantially cylindrical and extend longitudinally therethrough. The tubes 68 contain the powdered alkaloid composition 76.

[34] As shown in FIG. 2, a disposable multi-dose drum 66 may include hollow tubes 70 which have a diameter that is larger than the diameter of tubes 68. The diameter of each tube 68 is dependent on the volume and weight of dry powder to be delivered. Both tubes 68 and 70 may be packed within the rotating drum 66 so as to maximize the amount of doses available per rotating drum. For each tube 68, there is a corresponding hollow tube 70. One arrangement is for the powder-filled tubes 68 and hollow tubes 70 to be arranged in pairs vertical to each other. The drum 66 may be placed on spindle (not illustrated) which is inserted into a bore 72 such that the drum 66 is coaxial with the spindle. A clear sealed plastic overlay 86 may be disposed on the front and rear surfaces of the drum 66 to cover the tubes 68.

[35] A diffuser 112 includes an impact ball 114 at a portion that is proximate to the gas passage. The impact ball 114 is used to reduce the initial high velocity of highly turbulent gas and drug that enters diffuser 112. When gas and the alkaloid composition is injected into the diffuser 112, a high-energy flow may concentrate in the center of the unit. The impact ball 114 helps avoid this channeling effect. The diffuser 112 is shaped as an expansion cone to slow down the gas-powder mixture. The spacer 96 may be combined with the diffuser 112 so that the large particles can drop out of particle cloud in the spacer 96. Other details of this type of inhaler and variations thereof are described in U.S. Patent 7,461,649 to Gamard et al., the disclosure of which is hereby incorporated by reference in its entirety.

[36] Another, non- limiting example of an inhaler that may be used is shown in FIG. 3.

The inhaler of this embodiment is a one-piece tube 10 of generally uniform wall thickness. The tube 10 may be extruded, molded or fabricated out of flat stock such as a thermoplastic material having suitable gas barrier properties, and then rolled and sealed into a formed tube. The tube should be impermeable to the active agent(s). The tube 10 may have, for example, a wall thickness of about 0.002" to about 0.020". The tube 10 may be formed in a circular (as shown) or noncircular cross-section. The extruded tube may be cut into lengths between about 1" to 5" to resemble the length of conventional cigarettes. A plurality (e.g., 20) of the cut tubes may be provided in a pack (not illustrated) similar to a conventional cigarette pack.

[37] Once cut, the tube 10 is loaded with ingredients comprising the volatile delivery system. An absorbent material 12 compatible with the volatile agent(s) may be inserted into the tube 10 for delivery to the user. The absorbent material 12 can be in the form of a plug, strips, or the like. A liquid alkaloid composition (e.g., anatabine in an organic solvent) may be loaded into the tube 10 via spray or direct injection. The alkaloid composition may be loaded into the tube in a gaseous atmosphere, such as nitrogen or other gas, conducive to preserving the alkaloid(s). The tube 10 then may be pinched closed at the two open ends 16 and heat-sealed according to well known techniques. The heat seal affected at the two ends 16 prevents ambient air from entering the tube 10 or the ingredients within the tube from escaping to the atmosphere. Other details of this type of inhaler are shown in U.S. Patent 6,769,436 to Horian, the disclosure of which is hereby incorporated by reference in its entirety.

[38] By providing a balanced alkaloid composition containing anatabine as a significant alkaloid component, it is possible to prepare inhalers that reduce cravings for traditional tobacco smoking, while minimizing toxicity and other undesirable side effects associated with nicotine and other tobacco components. The inhaler may be used as needed to satisfy cravings, or at intervals such as once daily, twice daily, or three or more times daily, depending on such factors as the concentration of active components and the subject's physiological conditions.

EXAMPLES 1-16

[39] Alkaloid compositions for inhalers may be prepared by combining the components listed in Tables 1 and 2 below. Powdered compositions (Table 1) may be prepared by blending the alkaloid(s), excipient carriers, and flavors. Liquid compositions (Table 2) may be prepared by blending the alkaloid(s), solvents, water, and flavors. Anatabine may be prepared synthetically as described in Examples 1-3 of U.S. Patent 8,207,346 Bl . Nicotine, anabasine, and nornicotine may be extracted from tobacco materials and purified using known techniques.

Table 1

Example 1 2 3 4 5 6 7 8

Anatabine (mg) 10 8 4 6 7 12 8 6

Nicotine (mg) - 4 8 4 5 6 - 4

Nornicotine (mg) - - - - - - 4 2

Anabasine (mg) - - - 3 1 1 - 2

Mannitol (mg) 10 10 10 10 10 10 10 10

Flavors (g) 1 1.25 1.5 1 1 1.5 0.75 1.25 Table 2

EXAMPLE 17

[40] A liquid alkaloid composition for an inhaler is prepared by combining polyethylene glycol 400, anatabine (10 mg/ 10 mL) and nicotine to yield a composition containing 10 mg/ 10 mL of anatabine and 18 mg/ 10 mL of nicotine.

[41] The compositions described in Examples 1-17 may be filled into refilling cartridges for inhalers, or filled into a container that is used as part of a kit for refilling receptacles in inhalers. The compositions alternatively may be filled into a single-use or disposable type of inhaler.

EXAMPLE 18

[42] This example describes a medication-based treatment of nicotine addiction in a nonhuman primate model of nicotine self-administration. The acute effects of anatabine (0.18-3.2 mg/kg, IM) or saline on nicotine- and food-maintained responding were examined in seven rhesus monkeys. Nicotine (0.01 mg/kg/inj, base) and banana-flavored food pellets (lg) were available under a second-order schedule (FR 2 [VR 16:S]). Anatabine or saline injections were administered 15 min. before the midday food self-administration session began. Saline control treatment was in effect after administration of each anatabine dose.

[43] Seven adult rhesus monkeys (Macaca Mulatta) were trained to self-administer nicotine (0.01 mg/kg/inj [base]) and 1 g banana- flavored food pellets on a second- order schedule of reinforcement (FR 2 [VR 16:S]). Food was available during three 1-hour sessions each day (7 AM, 3 PM and 7 PM). A test session began at 11 ::00 AM and consisted of a 30 min food session and a 90 min nicotine session. A second nicotine session began immediately following the food session at 3 PM.

[44] A time-out period during which responding had no scheduled consequences followed each food and nicotine self-administration session. Nicotine injections were not limited during the 90 min test session, but were limited to 20 during the one-hour session. Food pellets were limited to 25 per session or 100 per day. The sequence of food and nicotine sessions is shown in the diagram. At the time of these studies, monkeys were nicotine-experienced and had at least five months of nicotine exposure.

[45] During test sessions, a single dose of anatabine (0.18-3.2 mg/kg, IM) or saline control treatment was given 15 min before the mid-day food self-administration session began. Following each dose of anatabine or saline, monkeys returned to stable baseline levels of nicotine- and food-maintained responding before the next treatment dose was administered. This control is essential to avoid confounding effects of the previous medication dose and to establish that catheter malfunction did not account for any decreases in nicotine self-administration.

[46] Systematic assessments to monitor any changes in behavior (e.g., sedation or agitation) were conducted after each saline or anatabine test session (Kato and Yanagita, 1981). Nicotine and anatabine solutions were prepared in sterile saline or cyclodextrin and filter- sterilized using a 0.22 μιη Millipore filter. Nicotine hydrogen tartrate was buffered with NaOH to achieve a pH of 6-7. Nicotine doses are expressed as the base.

[47] During saline control treatment monkeys earned an average of 20.37 ± 2.45 nicotine injections (0.01 mg/kg/inj) and 25 food pellets during the nicotine and food test sessions conducted at midday. Administration of single doses of anatabine (0.18 -3.2 mg/kg, IM) dose-dependently decreased nicotine self-administration at the training dose (0.01 mg/kg/inj [base]) (Figure 4). Decreases in nicotine-maintained responding were significantly different from baseline at anatabine doses of 1.8 and 3.2 mg/kg, IM (P<0.005-0.02). Food-maintained responding also decreased as a function of increasing doses of anatabine but these changes were not statistically significant.

[48] Subsequently, the nicotine dose-effect curve (0.001-0.1 mg/kg/inj) was determined under the same conditions in 7 monkeys (FIG. 5). The peak of the dose effect curve was at 0.0032 mg/kg/inj. Increasing doses of nicotine did not change food-maintained responding. Data for two monkeys suggest that a moderate dose of anatabine (0.32 mg/kg) shifts the peak of the dose-effect curve to the right. A higher dose of anatabine (1.0 mg/kg) flattened the dose-effect curve. Food-maintained responding also decreased at the peak reinforcing doses of nicotine (0.0032 and 0.01 mg/kg/inj) and there was considerable variability across animals.

[49] Anatabine was found to dose-dependently reduced nicotine self-administration (P<0.005-0.02) with no significant effects on food-maintained responding. Systematic behavioral assessments following each treatment session revealed no evidence of sedation or agitation that could disrupt operant responding. Each monkey returned to baseline levels of nicotine self-administration before administration of the next dose of anatabine, so catheter malfunction could not account for the significant decreases in nicotine self-administration observed. These findings in rhesus monkey are consistent with anatabine's effects on nicotine self-administration in rats (Caine et al., 2013). Anatabine (1.8-5.6 mg/kg) significantly reduced nicotine -maintained responding at doses of 0.003 and 0.01 mg/kg/inj (P<0.5-0.001) and flattened the nicotine dose-effect curve.

[50] FIG. 6 shows similar effects of anatabine on nicotine self-administration by rats.

Anatabine (1.8 and 3.2 mg/kg, IP) significantly reduced nicotine-maintained responding at peak reinforcing doses of 0.003 and 0.01 mg/kg/inj (P<0.05-0.001).

EXAMPLE 19

[51] This example describes a study conducted on anatabine along with other common nicotinic alkaloids in rat frontal and hippocampal homogenates. Table 3 shows binding data of anatabine relative to nicotine. Table 3

[52] A study was conducted to evaluate the in vitro effects of anatabine on three cloned human nicotinic acetylcholine receptor (nAChR) channels expressed in mammalian cells using a Fluo-8 calcium kit and a Fluorescence Imaging Plate Reader (FLIPRTETRA™) instrument. In this study, the ability of anatabine racemate to act as an agonist, positive allosteric modulator (PAM), or antagonist of the α3/β4, α4/β2 and α7 nAChR channels was tested. Nicotine (-) isomer was included as a comparator, and the endogenous neurotransmitter acetylcholine was included as a positive control.

[53] The results showed that anatabine functioned as an agonist at all three nAChR channels tested, but showed no antagonist or PAM effects at any of the receptor subtypes. In the agonist assay, the maximum level of stimulation for the α4β2 receptor was similar for anatabine and nicotine, indicating that anatabine acted as a full agonist at this receptor. However, the EC50 for anatabine 's agonist effects at the nAChR α4β2 receptor was approximately 200-fold less potent than nicotine (282 μΜ and 1.302 μΜ for anatabine and nicotine, respectively). For the α3β4 channel, the EC50 of nicotine was determined to be 9.37 μΜ and that of anatabine was 14.58 μΜ. The maximal response of the channel to nicotine approached that of ACh indicating that nicotine is a full agonist of the α3β4 receptor, as expected. However, the maximal response to anatabine was about 5 -fold lower than that of ACh indicating that anatabine is a partial agonist of this receptor. For the a7 assay, nicotine and anatabine were both potent agonists, and the EC50's were calculated to be approximately 0.008 μΜ for each compound. The maximum level of activation for both nicotine and activation was also similar (and similar to ACh), suggesting that anatabine and nicotine are full, potent agonists of the a7 nAChR. [54] To summarize, the results of this study showed that anatabine is an agonist of all three nAChR channels tested, but with different characteristics at each subtype, and with some significant differences relative to nicotine. Although anatabine was a full agonist at the α4β2 receptor, it was much less potent than nicotine or ACh. Anatabine showed partial agonist activity towards the α3β4 receptor, and was only slightly less potent than nicotine. Finally, anatabine showed full agonist activity at the a7 nAChR channel, with similar potency as nicotine.

[55] These data indicate that anatabine may activate α4β2 receptors, but very high concentrations beyond any recommended or tolerated doses are likely needed to produce significant agonist effects. This suggestion is supported by behavioral data which showed that rodents trained to self-administer nicotine do not find anatabine to be rewarding, and that anatabine administration does not reverse precipitated nicotine withdrawal. Nicotine is the prototypic full α4β2 agonist, and its higher potency at this receptor is likely at least partially responsible for the reinforcing and dependence- producing effects of tobacco products. Conversely, anatabine was shown to be a full and highly potent agonist at the a7 receptor, suggesting that this may be one mechanism by which anatabine exerts anti-inflammatory effects such as those that have been observed in both in vitro and in vivo studies. Although it is not selective for the a7 receptor, the findings from this study suggest that anatabine 's activity toward this particular nAChR subtype is similar to nicotine, and provide evidence that this may be one pathway through which anatabine 's immuno-modulatory effects are mediated.

[56] While the invention has been described with respect to specific examples, those skilled in the art will appreciate that there are numerous variations and permutations of the above described systems and techniques that fall within the spirit and scope of the invention as set forth in the appended claims.