Patent Application
20050009903
1. Field of the Invention
The invention generally relates to cannabinoid analogues. In particular, the invention provides cannabinoid analogues that exhibit specificity for the CB2 cannabinoid receptor.
2. Background of the Invention
The complex effects of cannabinoids are considered to be mediated through at least two G-protein coupled, transmembrane receptors. One of these, designated as CB1, is found predominantly in the central nervous system and is responsible for most of the overt pharmacological effects of cannabinoids. A second receptor, designated CB2 was originally identified from macrophages present in the spleen, and is expressed primarily in the periphery. Although it has been known for some time that cannabinoids are involved in immunomodulation, the discovery that CB2 receptors are expressed primarily in the immune system led to the suggestion that the CB2 receptor was responsible for the immunomodulatory effects of cannabinoids. This suggestion has recently been confirmed by the observation that such effects are absent in CB2 receptor knockout mice. Furthermore, recent studies have now highlighted other beneficial effects specific to the CB2 receptor. These effects include the discovery that the CB2 selective ligand, JWH-133, is effective in reducing spasticity in the mouse model of multiple sclerosis. More pertinently, the same CB2 selective ligand inhibits the in vivo growth of glioma tumors. Other effects modulated by the CB2 receptor include peripheral antinociception and, at least in part, the antitumor properties of ajulemic acid.
There is an ongoing need for the development of ligands which are specific for the CB2 receptor.
It is an object of this invention to provide cannabinoid analogues which bind preferentially to the CB2 cannabinoid receptor, and which exhibit little or no binding affinity for the CB1 receptor. The analogues are 1-methoxy-, 1-deoxy-11-hydroxy- and 11-hydroxy-1-methoxy-Δ8-tetrahydrocannabinols and 1-alkyl-3(1-naphthoyl)indoles.
In one embodiment, the invention provides a compound of general formula
wherein R1 and R2 are H, OH or OCH3; n ranges from 0 to 4; and R1 and R2 are not both H. A steric representation of this general formula is
where R1 is H or OH; R2 is H or OCH3; n ranges from 0 to 4; and R1 and R2 are not both H. In one embodiment of the compound, R1 is H and R2 is OCH3. In another embodiment, R1 is OH and R2 is H. In yet another embodiment, R1 is OH and R2 is OCH3.
The invention further provides a method for selectively binding to CB2 receptors and not to CB1 receptors. The method comprises the step of exposing the CB2 receptors and the CB1 receptors to a compound of general formula
wherein R1 and R2 are H, OH, or OCH3; n ranges from 0 to 4; and R1 and R2 are not both H. One possible steric representation of the general formula is
where R1 is H or OH; R2 is H or OCH3; n ranges from 0 to 4; and R1 and R2 are not both H. In preferred embodiments: R1 is H and R2 is OCH3; or R1 is OH and R2 is H; or R1 is OH and R2 is OCH3.
In one embodiment of the invention, the compound binds to the CB2 receptors in an amount sufficient to treat cancer, for example, a glioma tumor. In another embodiment, the compound binds to the CB2 receptors in an amount sufficient to treat pain, e.g. inflammatory pain.
The present invention provides a method for killing tumor cells. The method includes the step of exposing the tumor cells to a compound of general formula
wherein R1 and R2 are H, OH, or OCH3; n ranges from 0 to 4; and R1 and R2 are not both H. One possible steric representation of the general formula is
where R1 is H or OH; R2 is H or OCH3; n ranges from 0 to 4; and R1 and R2 are not both H. In preferred embodiments: R1 is H and R2 is OCH3; or R1 is OH and R2 is H; or R1 is OH and R2 is OCH3. The tumor cells may be glioma tumor cells.
The invention further provides a compound of general formula
wherein R1, R2, R3, R4 and R5 are H, CH3, or OCH3; and n ranges from 1 to 5. In some embodiments of the invention: R1 is H or CH3; R2 is H or OCH3; R3 is H, CH3, or OCH3; R4 is H or OCH3; and R5 is H or CH3. Examples of such compounds include those where: 1) n is 3; R1, R3, R4 and R5 are H; and R2 is OCH3; 2) n is 3; R1 is CH3; R2 is OCH3; and R3, R4 and R5 are H; 3) n is 5; R1, R3, R4 and R5 are H; and R2 is OCH3; 4) n is 5; R1 is CH3, R2 is OCH3 and R3, R4 and R5 are H; 5) n is 3 or 5; R1, R2, R3 and R5 are H; and R4 is OCH3; 6) n is 3 or 5; R1 is CH3; R2, R3 and R5 are H; and R4 is OCH3; 7) n is 3; R1, R2, R4 and R5 are H; and R3 is CH3; and 8) n is 3 or 5; R1 is CH3;R2, R3 and R4 are H; and R5 is CH3.
The invention also provides a method for selectively binding toCB2 receptors and not to CB1 receptors. The method includes the step of exposing the CB2 receptors and the CB1 receptors to a compound of general formula
where R1, R2, R3, R4 and R5 are H, CH3, or OCH3; and n ranges from 1 to 5. In some embodiments: R1 is H or CH3; R2 is H or OCH3;R3 is H, CH3, or OCH3; R4 is H or OCH3; and R5 is H or CH3. In preferred embodiments: 1) n is 3; R1, R3,R4 and R5 are H; and R2 is OCH3; 2) n is 3; R1 is CH3;R2 is OCH3; and R3, R4 and R5 are H; 3) n is 5; R1, R3, R4 and R5 are H; and R2 is OCH3; 4) n is 5; R1 is CH3, R2 is OCH3 and R3, R4 and R5 are H; 5) n is 3 or 5; R1, R2, R3 and R5 are H; and R4 is OCH3; 6) n is 3 or 5; R1 is CH3; R2, R3 and R5 are H; and R4 is OCH3; 7) n is 3; R1, R2, R4 and R5 are H; and R3 is CH3; and 8) n is 3 or 5; R1 is CH3; R2, R3 and R4 are H; and R5 is CH3.
The compound binds to the CB2 receptors in an amount sufficient to treat cancer, e.g. a glioma tumor. Alternatively, the compound binds to the CB2 receptors in an amount sufficient to treat pain, e.g. inflammatory pain.
The present invention also provides a method for killing tumor cells. The method includes the step of exposing the tumor cells to a compound of general formula
where R1, R2, R3, R4 and R5 are H, CH3, or OCH3; and n ranges from 1 to 5.
In some embodiments: R1 is H or CH3; R2 is H or OCH3;R3 is H, CH3, or OCH3; R4 is H or OCH3; and R5 is H or CH3. In preferred embodiments: 1) n is 3; R1, R3, R4 and R5 are H; and R2 is OCH3; 2) n is 3; R1 is CH3; R2 is OCH3; and R3, R4 and R5 are H; 3) n is 5; R1, R3, R4 and R5 are H; and R2 is OCH3; 4) n is 5; R1 is CH3, R2 is OCH3 and R3, R4 and R5 are H; 5) n is 3 or 5; R1, R2, R3 and R5 are H; and R4 is OCH3; 6) n is 3 or 5; R1 is CH3; R2, R3 and R5 are H; and R4 is OCH3; 7) n is 3; R1, R2, R4 and R5 are H; and R3 is CH3; and 8) n is 3 or 5; R1 is CH3; R2, R3 and R4 are H; and R5 is CH3.
In a preferred embodiment, the tumor cells are glioma tumor cells.
The present invention provides compounds that bind with high affinity to the CB2 cannabinoid receptor and methods for their use. By binding with “high affinity” we mean that the Ki values of the compounds for the CB2 receptor are in the range of about 0.03 nM to about 40 nM. In a preferred embodiment, the compounds display both high binding affinity for the CB2 receptor and high binding selectivity for the CB2 receptor. By “binding selectivity” we mean that the compounds exhibit an affinity for the CB2 cannabinoid receptor that is in the range of about 10 to about 200 fold or more higher than their affinity for the CB1 cannabinoid receptors. In other words, the Ki values of the compounds for CB2 receptors is in the range of about 10 to about 200 times or more lower than the Ki values of the compounds for the CB1 receptor. Measurement of the Ki values may be carried out in any of several known assay systems, including but not limited to biological tissue such as brain and spleen, or in cells that are genetically modified to express CB1 or CB2 receptors. In a preferred embodiment, the assay system is brain tissue that contains CB1 receptors and cells genetically modified to express CB2 receptors.
In one embodiment of the invention, the compounds are of the general formula represented in Formula 1 below.
In Formula 1, R1 and R2 may be H, OH or OCH3, but R1 and R2 are not both H; and n may range from 0 to 4. In some embodiments of the invention, R1 is H or OH and R2 is H or OCH3, In preferred embodiments of the invention:
In another embodiment of the invention, the compounds of the invention are of the general formula represented in Formula 2 below.
In Formula 2, R1, R2, R3, R4 and R5 are H, CH3, or OCH3; and n ranges from 1 to about 5. In some embodiments, R1 is H or CH3; R2 is H or OCH3; R3 is H, CH3, or OCH3; R4 is H or OCH3; and R5 is H or CH3. In preferred embodiments of the invention: n is 3 or 5, R1 is H or CH3, R2 is OCH3 and R4 and R5 are H (e.g. compounds 30-33 of Example 2, listed in Table 3, where:
The compounds of the present invention may be useful as analgesics in a variety of applications including but not limited to pain management. Pain of any type may be ameliorated by the administration of the compounds of the present invention. In preferred embodiments, the compounds of the present invention are administered to treat acute and chronic inflammatory pain (e.g., inflammation from autoimmune disorders such as arthritis, inflammation from injuries or from surgery, etc.).
In addition, the compounds of the present invention may be used for the treatment of cancer. Examples of types of cancer that may be treated by the administration of the present compounds include but are not limited to glioma tumors and non-melanoma skin cancers. In a preferred embodiment, the compounds are used to treat glioma tumors.
The invention thus provides a method for treating cancer in a patient in need thereof. By “treating” cancer we mean that the compound is administered in order to alleviate symptoms of the disease (e.g. to decrease tumor size, halt metastasis, etc.). Those of skill in the art will recognize that, in the treatment of cancer, administration of the compounds of the present invention may completely eradicate symptoms of the disease, or, alternatively, may attenuate or slow the progression of disease, which is also beneficial to the patient.
While in preferred embodiments, the compounds of the present invention may be used to treat inflammatory pain and cancer, in a more general sense, the invention further provides methods of treating conditions or disorders related to CB2 receptor-regulated systems. By “CB2receptor-regulated systems” we mean biochemical pathways which include the binding of a cannabinoid to the CB2 receptor molecule within the pathway. Examples of such biochemical pathways include but are not limited to activation of G-proteins and subsequent inhibition of adenylyl cyclase. The compounds of the present invention may be used to treat aberrations in CB2 receptor-regulated systems, or to alter CB2 receptor-regulated systems when it is desirable to do so in order to ease untoward symptoms in a patient. For example, patients with pain or spasticity arising from inflammation or neuropathological damage will benefit from activation of the CB2 receptor with a selective agonist. In addition, the compounds may also be used for research purposes.
Implementation of the claimed invention will generally involve identifying patients suffering from the indicated disorders and administering the compounds of the present invention in an acceptable form by an appropriate route. The exact dosage to be administered may vary depending on the age, gender, weight and overall health status of the individual patient, as well as the precise etiology of the disease. However, in general for administration in mammals (e.g. humans), dosages in the range of from about 0.1 to about 30 mg of compound per kg of body weight per 24 hr., and more preferably about 0.1 to about 10 mg of compound per kg of body weight per 24 hr., are effective.
Administration may be oral or parenteral, including intravenously, intramuscularly, subcutaneously, intradermal injection, intraperitoneal injection, etc., or by other routes (e.g. transdermal, sublingual, oral, rectal and buccal delivery, inhalation of an aerosol, etc.).
The compounds may be administered in the pure form or in a pharmaceutically acceptable formulation including suitable elixirs, binders, and the like (generally referred to a “carriers”) or as pharmaceutically acceptable salts (e.g. alkali metal salts such as sodium, potassium, calcium or lithium salts, ammonium, etc.) or other complexes. It should be understood that the pharmaceutically acceptable formulations include liquid and solid materials conventionally utilized to prepare both injectable dosage forms and solid dosage forms such as tablets and capsules and aerosolized dosage forms. In addition, the compounds may be formulated with aqueous or oil based vehicles. Water may be used as the carrier for the preparation of compositions (e.g. injectable compositions) which may also include conventional buffers and agents to render the composition isotonic. Other potential additives and other materials (preferably those which are generally regarded as safe [GRAS]) include: colorants; flavorings; surfactants (TWEEN, oleic acid, etc.); solvents, elixirs, and binders or encapsulants (lactose, liposomes, etc). Solid diluents and excipients include lactose, starch, conventional disintergrating agents, coatings and the like. Preservatives such as methyl paraben or benzalkium chloride may also be used. Depending on the formulation, it is expected that the active composition will consist of about 1% to about 99% of the composition and the vehicular “carrier” will constitute about 1% to about 99% of the composition. The pharmaceutical compositions of the present invention may include any suitable pharmaceutically acceptable additives or adjuncts to the extent that they do not hinder or interfere with the therapeutic effect of the active compound.
The administration of the compounds of the present invention may be intermittent, or at a gradual or continuous, constant or controlled rate to a patient. In addition, the time of day and the number of times per day that the pharmaceutical formulation is administered may vary are and best determined by a skilled practitioner such as a physician. Further, the effective dose can vary depending upon factors such as the mode of delivery, gender, age, and other conditions of the patient, as well as the extent or progression of the disease. The compounds may be provided alone, in a mixture containing two or more of the compounds, or in combination with other medications or treatment modalities. The compounds may also be added to blood ex vivo and then be provided to the patient.
The complex pharmacological effects of cannabinoids are considered to be mediated through at least two G-protein-coupled, transmembrane receptors. One of these, designated as CB1, is found predominantly in the central nervous system and is responsible for most of the overt pharmacological effects of cannabinoids.1-4 A second receptor, designated CB2, was originally identified from macrophages present in the spleen, and is expressed primarily in the periphery.5 Very recently evidence has been presented for the existence of a third cannabinoid receptor, which has been detected in mouse brain.6 It is generally accepted that the CB1 receptor is implicated in eliciting the in vivo effects of cannabinoids; a good correlation has been found between the CB1 receptor affinities of a series of cannabinoids and their in vivo effects.7,8 These in vivo effects are blocked by SR141716A, an inverse agonist for the CB1 receptor, and are absent in CB1 receptor knockout mice.9,10
Although it has been known for some time that cannabinoids are involved in immunomodulation,11 the discovery that the CB2 receptor is expressed primarily in cells of the immune system led to the suggestion that the CB2 receptor was responsible for the immunomodulatroy effects of cannabinoids.5 This suggestion has been confirmed recently by the observation that these immunomodulatory effects are absent in CB2 receptor knockout mice.12 Although there is evidence that the CB2 receptor is not expressed in the central nervous system,13 it has recently been found that this receptor is expressed in adult rat retina.14
Although it has been known for several years that the CB2 receptor is expressed in cells in the immune system, it has only been within the past few years that specific effects mediated by this receptor have been recognized. These effects included the discovery that a CB2 selective receptor ligand, JWH-133, is effective in reducing spasticity in the mouse model of multiple sclerosis,15 and the same CB2 selective ligand also inhibits the in vivo growth of glioma tumors.16 Other effects modulated by the CB2 receptor include peripheral antinociception,17 and at least in part, the antitumor properties of ajulemic acid.18
Several years ago we reported that 3-(1′,1′-dimethyl-heptyl)-1-deoxy-11-hydroxy-668-tetrahydrocannabinol (1-deoxy-11-hydroxy-Δ8-THC-DMH,deoxy-HU-210,JWH-051 (1 in
Others have described two 1-methoxy cannabinoids,1-methoxy-Δ8-THC-DMH (4 in
In view of the continuing recognition of the importance of the CB2 receptor, we have taken advantage of the currently available knowledge of the SAR of 1-methoxy-and 1-deoxy-668-THC analogues to design three series of CB2 selective cannabinoid receptor ligands. Based on the knowledge that the side chain in the 1-deoxy-Δ8-THC series can be shortened significantly without seriously attenuating CB2 receptor affinity, and that an 11-hydroxyl enhances both CB1 and CB2 receptor affinities,20 the initial synthetic targets included a series of 1-deoxy-3-(10,10-dimethylalkyl)-11-hydroxy-Δ8-THC analogues. Since 1-methoxy-Δ8-THC-DMH (4) is also a CB2 selective cannabinoid ligand,20,21,23 a series of 3-(1′,1′-dimethylalkyl)-1-methoxy-Δ8-THCs was also prepared. The third series of CB2 selective compounds consisted of 11-hydroxy-3-(1′,1′-dimethylalkyl)-1-meth-oxy-Δ8-THC analogues which combined the structural features of the other two series.
Results
The 1-methoxy-Δ8-THC analogues (6a-6e,
The 1-deoxy-11-hydroxy-Δ8-THCs (9a-9e,
In the 11-hydroxy-3-(1′,1′-dimethylalkyl)-1-methoxy-Δ8-THC series, in addition to the dimethylethyl through dimethylhexyl analogues (11a-11e,
The affinities of 1-methoxy-, 11-hydroxy-and 11-hydroxy-1-methoxy-Δ8-THC analogues 6 (
aref 8;
bref 30;
cref 20;
dref 19.
In the 1-methoxy-Δ8-THC series (6a-6e,
As mentioned above, our CB1 receptor affinities were determined using a rat brain membrane preparation while Gareau et al. employed a human CB1 receptor preparation which was not described in detail.21 The binding assays described by Ross et al. were carried out using CHO (Chinese hamster ovary) cells transfected with human CB1 and CB2 receptors.23 Our CB2 data were obtained as described in the Experimental using HEK (human embryonic kidney) cells transfected with human CB2 receptors.
The 1-deoxy-11-hydroxy-Δ8-THC analogues (9a-9e,
The compounds in the 11-hydroxy-3-(1′,1′-dimethylalkyl)-1-methoxy-Δ8-THC series (11a-11f,
The data summarized in Table 1, are in general agreement with the preliminary SAR for the CB2 receptor which we developed based upon our study of 1-deox-Δ8-THC analogues.20 In the 1′,1′-dimethyl-1-deoxy-Δ8-THC series described previously, those compounds with a three to seven carbon side chain (2 and 8b-8e) all have high affinity for the CB2 receptor (Ki<20 nM).Of the three new series of CB2 selective cannabinoid receptor ligands, only the 1-deoxy-11-hydroxy-Δ8-THC analogues (1 and 9a-9e) show uniformly high affinity for the CB2 receptor, with Ki=0.032±0.019 nM for the dimethyl-heptyl analogue (1)19 to Ki=18.1±1.8 nM for the lowest member of the homologous series (9a). There is a progressive improvement in CB2 receptor affinity as the length of the side chain increases from two to seven carbon atoms. These compounds also show from modest to high affinity for the CB1 receptor, increasing from Ki=270±58 nM for the dimethylethyl analogue (9a) to Ki=1.2±0.1 nM for the dimethylheptyl compound (1) reported previously.19 The relatively high CB1 receptor affinities for the compounds in this series may be attributed to the 11-hydroxyl group serving as a surrogate for the phenolic hydroxyl in more traditional cannabinoids as suggested by molecular modeling studies carried out on 1, combined with a 3-(1′,1′-dimethyl-alkyl) substituent of sufficient length to interact with the lipophilic portion of the receptor.19
The compounds in the 1-methoxy series (4 and 6a to 6e) all have little affinity for the CB1 receptor, with CB2 affinities ranging from very slight for the dimethylethyl analogue (6a, Ki=1867±867 nM) to quite high for the dimethylhexyl compound (6e, Ki=18±2 nM). As reported previously, the dimethylheptyl analogue (4) has little affinity for the CB1 receptor and moderate affinity for the CB2 receptor. The dimethylhexyl methyl ether (6e) is a highly selective CB2 receptor ligand with good affinity for the CB2 receptor and very little affinity for the CB1 receptor.
The compounds of the 11-hydroxy-1-methoxy series (11a-11f) are intermediate between those of the other two series of ligands in their affinities for both receptors. The lower members of this series (11a-11c) have little affinity for the CB1 receptor with Ki=1856±148 nM for the dimethylethyl analogue (11a) and Ki=347±34 mM for the dimethylbutyl compound (11c). The higher members of this series have from moderate affinity (11d, Ki=40±6 nM) for the CB2 receptor to high affinity for the dimethylhexyl (11e) and dimethylheptyl (11f) analogues. The affinities of 11e and 11f are identical within experimental error for each receptor, with Ki=14 nM at CB1 and 1.2 nM at CB2.
In terms of the SAR for 1-methoxy-, 1-deoxy-11-hydroxy and 11-hydroxy-1-methoxy-Δ8-THC analogues, it is apparent that an 11-hydroxyl substituent enhances affinity for both the CB1 and CB2 receptors. Also, in the 3-(1,1-dimethylalkyl) series the length of the side chain plays a critical role in determining affinity for both receptors. It is somewhat important for CB2 affinity, particularly in the methyl ether series (6),but for significant CB1 affinity a chain length of at least five carbon atoms is essential.
In summary, although several of these compounds show selectivity for the CB2 receptor, only five of them, 1-methoxy cannabinoids 6d and 6e, 1-deoxy-11-hydroxy compounds 9a and 9b, and 11-hydroxy-3-(1′,1′-dimethylbutyl)-1-methoxy-Δ8-THC (11c) have a combination of high affinity for the CB2 receptor and little affinity for the CB1receptor. Only 3-(1′,1′-dimethyl-hexyl)-1-methoxy-Δ8-THC (6e, JWH-229) with Ki=3134±110 nM at CB1 and Ki=18±2 nM at CB2 is comparable in selectivity to 1-deoxy-3-(1′,1′-dimethylhexyl)-Δ8-THC (3, JWH-133) with Ki=677±132 nM at CB1 and Ki=3.4±1.0 nM at CB2.20 Although JWH-229 (6e) has slightly less affinity for the CB2 receptor than JWH-133 (3), it has significantly lower affinity for CB1, and is thus a potentially useful CB2 selective cannabinoid ligand with very little affinity for CB1.
Experimental
General
IR spectra were obtained using Nicolet 5DX or Magna spectrometers; 1H and 13C NMR spectra were recorded on a Bruker 300AC spectrometer. Mass spectral analyses were performed on a Hewlett-Packard 5890A capillary gas chromatograph equipped with a mass sensitive detector. HRMS data were obtained in the Mass Spectrometry Laboratory, School of Chemical Sciences, University of Illinois. Ether and THF were distilled from Na-benzophenone ketyl immediately before use, and other solvents were purified using standard procedures. Column chromatography was carried out on Sorbent Technologies silica gel (32-63 μm) using the indicated solvents as eluents. All new compounds were homogeneous to TLC and 13C NMR. All target compounds were homogeneous to GLC or TLC in two different solvent systems. TLC was carried out using 200 m silica gel plates using the indicated solvents. GLC analyses were performed on the Hewlett-Packard 5890A GC/MS using a 60 m carbowax column and helium gas as a carrier. An initial column temperature of 60° C. was employed and the temperature was increased at a rate of 1.5 C/min to a maximum temperature of 300° C. with a total run time of 20 min. Elemental analyses were performed by Atlantic Microlab, Norcross, Ga.
1-Methoxy-3-(1′,1′-dimethylpropyl)-Δ8-THC (6b).To a solution of 0.569 g (1.8 mmol) of 3-(1′,1′-dimethylpropyl)-Δ8-THC (7b)20 in 14 mL of dry DMF under N2 was added 0.151 g (2.7 mmol) of KOH and 0.33 mL (5.4 mmol)of methyl iodide. The reaction mixture was stirred for 48 h at ambient temperature before being quenched by the addition of 2 mL of aqueous NH4Cl and removal of the DMF in vacuo. The residue was extracted with three portions of ether and the combined organic extracts were dried (MgSO4) and concentrated in vacuo. The crude product was initially purified by dry flash chromatography (petroleum ether:ether, 97:3) followed by gradient elution chromatography (petroleum ether:dichloromethane, 9:1 to 8:1)to afford 0.309 g (52%)of 6b as a colorless oil. Further chromatography (petroleum ether:dichloromethane, 85:15) of 0.104 g of this material gave 0.100 g of pure 6b, Rf 0.46 (petroleum ether:dichloromethane, 85:15); 1H NMR (500 MHz, CDCl3) δ 0.79 (t, J=7.1 Hz,3H),1.09 (s,3H),1.24 (s, 6H),1.38 (s,3H),1.54.1.60 (m,2H),1.70 (s,3H),1.74. 1.81 (m,3H),2.12.2.14 (m,1H),2.67 (td, J=4.7, 11.1 Hz,1H),3.15 (dd, J=4.7, 17.1 Hz,1H),3.81 (s, 3H),5.42 (br.s,1H),6.38 (d, J=1.6 Hz,1H),6.42(d, J=1.6 Hz,1H); 13C NMR (125.8 MHz,CDCl3) δ 14.1, 18.4, 23.4, 27.8, 27.9, 28.7, 31.7, 36.3, 37.6, 44.3, 45.0, 55.1, 76.4, 100.7, 108.3, 111.2, 119.2, 135.0, 149.8, 153.9, 158.6; MS (ED) m/z 328 (65), 299 (45), 245 (100); [αD20 −210° (c=0.29,CH2Cl2) HRMS calcd for C22H32O2: 328.2404, found 328.2402.
1-Methoxy-3-(1′,1′-dimethylethyl)-Δ8-THC (6a). Methoxy cannabinoid 6a was prepared by the procedure described above for the preparation of 6b. Methylation of 0.384 g (1.28 mmol) of 7a20 gave 0.173 g (43%)of 6a as a colorless oil: 1H NMR (300 MHz, CDCl3) δ 1.10 (s,3H), 1.29 (s,9H), 1.38 (s,3H), 1.74 (s,3H), 1.76.1.82 (m,3H), 2.11.2.14 (m,1H),2.65 (td, J=4.5, 10.9 Hz, 1H), 3.15 (dd, J=3.8, 17.3 Hz, 1H), 3.81 (s,3H), 5.41 (d, J=4.6 Hz,1H),6.44 (d, J=1.6 Hz, 1H), 6.48 (d, J=1.6 Hz, 1H); 13C NMR (75.5 MHz,CDCl3) δ 18.5, 23 (5), 27.6, 28.0, 31.2, 31.7, 34.7, 36.1, 45.0, 55.1, 76.5, 100.2, 107.6, 111.7, 119.2, 135.0, 150.9, 154.0, 158.7; MS (EI) m/z 314 (52), 299 (8), 246 (13), 231 (100);[αD20 −206° (c=2.65,CH2Cl2); HRMS calcd for C21H30O2: 314.2245, found 314.2246.
11-Methoxy-3-(1′,1′-dimethylbutyl)-Δ8-THC (6c). Methoxy cannabinoid 6c was prepared by the procedure described above for the preparation of 6b. Methylation of 0.782g (2.38 mmol)of 7c20 gave 0.461 g (43%)of 6c as a colorless oil: 1H NMR (500 MHz,CDCl3) δ 0.82(t, J=7.4 Hz, 3H), 1.09 (s, 3H), 1.05.1.15 (m, 2H), 1.25 (s, 6H),1.38 (s, 3H), 1.49.1.55 (m, 2H),1.70 (s, 3H), 1.74.1.85 (m, 3H), 2.09.2.16 (m, 1H), 2.66 (td, J=4.6, 11.0 Hz, 1H), 3.16 (dd, J=3.2, 16.5 Hz, 1H), 3.78 (s,3H), 5.42 (d,4.1H), 6.38 (d, J=0.9 Hz, 1H), 6.43 (d, J=0.9 Hz, 1H); 13C NMR (125.8 MHz,CDCl3) δ 14.9, 18.1, 18.5, 23.7, 27.7, 28.1, 28.9, 29.0, 31.8, 36.3, 37.8, 45.2, 47.2, 55.2, 76.9, 100.9, 108.3, 111.7, 119.4, 135.1, 149.7, 154.1, 158.8; MS (EI) m/z 342(37), 300 (100), 286 (20),259 (38); [αD20 −258° (c=0.79,CHCl3); HRMS calcd for C23H34O2: 342.2564, found 342.2559.
1-Methoxy-3-(1′,1′-dimethylpentyl)-Δ8-THC (6d). Methoxy cannabinoid 6d was prepared by the procedure described above for the preparation of 6b. Methylation of 2.80 g (8.17 mmol) of 7d20 gave 2.83 g (97%)of 6d as a colorless oil: 1H NMR (300 MHz,CDCl3) δ 0.83 (t, J=7.1 Hz, 3H), 1.03.1.11 (m, 2H), 1.10 (s, 3H), 1.16.1.29 (m,2H), 1.25 (s,6H), 1.38 (s,3H), 1.51.1.60 (m, 2H), 1.70 (s,3H), 1.73.1.90 (m,3H), 2.05.2.12 (m,1H), 2.66 (td, J=4.7, 10.8 Hz,1H), 3.06 (dd, J=4.0 Hz,1H), 3.81 (s,3H),5.41 (d, J=4.8 Hz,1H), 6.38 (d, J=1.6 Hz, 1H),6.43 (d, J=1.6 Hz,1H); 13C NMR (75.5 MHz, CDCl3) δ 14.1,18.4, 23.4,23.5, 26.9, 27.6, 27.9, 28.8, 31.7, 36.2, 37.6, 44.3, 45.0, 55.1, 76.5, 100.7, 108.2, 111.6, 119.2, 135.0, 149.7, 153.9, 158.7; MS (EI) m/z 356 (45), 300 (100), 286 (25), 273 (30); HRMS calcd for C24H36O2: 356.2716, found 356.2715; [αD20 −234° (c=0.24,CHCl3).
1-Methoxy-3-(1′,1′-dimethylhexyl)-Δ8-THC (6e). Methoxy cannabinoid 6e was prepared by the procedure described above for the preparation of 6b. Methylation of 2.59 g (7.26 mmol)of 7e20 gave 2.54 g (94%) of 6e as a colorless oil: 1H NMR (300 MHz,CDCl3) δ 0.83 (t, J=7.1 Hz, 3H), 1.01.1.30 (m,6H), 1.10 (s, 3H), 1.24 (s, 6H), 1.38 (s, 3H),1.48.1.56 (m,2H), 1.65.1.90 (m, 3H), 1.70 (s,3H), 2.11.2.18 (m,1H), 2.67 (td, J=4.7, 10.9 Hz, 1H), 3.15 (dd, J=4.1, 17.0 Hz, 1H), 3.81 (s, 3H), 5.41 (s, 1H), 6.38 (d, J=1.6 Hz, 1H), 6.42(d, J=1.6 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 14.1, 18.4, 22.5, 23.5, 24.3, 27.6, 27.7, 28.0, 28.8, 31.7, 32.6, 36.2, 37.7, 44.5, 45.1, 55.1, 76.4, 100.7, 108.3, 111.6, 119.2, 135.1, 149.7, 153.9, 158.7; MS (EI) m/z 370 (40), 300 (100), 287 (35); [αD20 −219° (c=0.21,CHCl3); HRMS calcd for C25H38O2: 370.2871, found 370.2872.
1-Deoxy-3-(1′,1′-dimethylpropyl)-11-hydroxy-Δ8-THC (9b).To a stirred suspension of 1.07 g (3.58 mmol) of 1-deoxycannabinoid 8b20 in 16 mL of ethanol at ambient temperature was added dropwise over 30 min a solution of 0.96 g of SeO2 (8.73 mmol) in 17.6 mL of ethanol/water (10:1). The reaction mixture was heated at reflux for 18 h, filtered through a pad of Celite, which was subsequently washed with three portions of methanol, and the combined organic extracts were concentrated in vacuo. The residue was extracted with three portions of ether and the resulting ethereal solution was washed successively with water then saturated NaHCO3.The organic phase was dried (MgSO4)and concentrated in vacuo to afford crude aldehyde 10b which was purified by dry flash chromatography (ethyl acetate:petroleum ether, 9:1) to afford 0.96 g of 10b as a light brown oil which was used without further purification. To a solution of 0.96 g (3.10 mmol) of the crude aldehyde in 35 mL of dry THF at 0° C. under N2was added 0.12 g (3.07 mmol) of LiAlH4.The reaction mixture was allowed to warm to room temperature, stirred for 2 h and then quenched with aqueous NH4Cl. After filtering through a pad of Celite, which was subsequently washed with diethyl ether, the combined organic extracts were dried (MgSO4)and concentrated in vacuo to afford the crude product. Initial chromatography (gradient elution with 17% diethyl ether to 35% diethyl ether in petroleum ether) gave a pale yellow resin which was further purified by chromatography (gradient elution with 5% acetone to 7% acetone in petroleum ether) to afford 0.31 g (32% for two steps)of 9b as a white foam: 1H NMR (500 MHz,CDCl3) δ 0.69 (t, J=7.8 Hz,3H), 1.17 (s,3H), 1.24 (s,6H), 1.40 (s,3H), 1.60 (q, J=7.4 Hz, 1H), 1.72.2.02 (m, 5H), 2.20.2.28 (m,1H), 2.69 (td, J=5.5, 11.0 Hz,1H), 2.80 (dd, J=4.2, 16.5 Hz,1H), 4.05 (d, J=12.8 Hz,1H), 4.08 (d, J=12.8 Hz, 1H), 5.76 (br.s, 1H), 6.76 (d, J=1.8 Hz, 1H), 6.85 (dd, J=1.8, 7.8 Hz, 1H), 7.14 (dd, J=0.9, 8.2 Hz, 1H); 13C NMR (125.8 MHz,CDCl3) δ 9.34, 19.2, 27.3, 27.8, 28.5, 31.9, 32.1, 36.9 ,37.7, 43.2, 67.0, 76.9, 114.9, 118.0, 121.7, 122.2, 126.2, 137.1, 149.4, 152.6; MS (EI) m/z 314 (59), 312 (20), 299 (13), 285 (100), 207 (100); [αD20 −126° (c=9.0,CHCl3); HRMS calcd for C21H30O2: 314.2251, found 314.2246.
1-Deoxy-3-(1′,1′-dimethylethyl)-11-hydroxy-Δ8-THC (9a). 11-Hydroxycannabinoid 9a was prepared by the procedure described above for the preparation of 9b. Stepwise oxidation and reduction of 1.27 g (4.46 mmol) of 8a20 gave 0.35 g (26% for two steps) of hydroxy cannabinoid 9a as a white foam: 1H NMR (500 MHz, CDCl3) δ 1.17 (s,3H), 1.29 (s,9H), 1.41 (s,3H), 1.76 dt, J=5.8, 13.0 Hz, 1H), 1.80 .1.90 (m, 1H), 1.94 (br. t, J=13.0 Hz, 1H), 2.08.2.28 (m,2H), 2.69 (dt, J=5.8, 3.1 Hz,1H), 2.80 (dd, J=5.3, 17.5 Hz,1H), 4.04 (d, J=17.0 Hz,1H), 4.08 (d, J=17.0 Hz,1H), 5.77 (br. s, 1H), 6.84 (d, J=1.4 Hz, 1H),6.92(dd, J=1.8, 8.2 Hz, 1H),7.17 (d, J=8.2 Hz, 1H); 13C NMR (125.8 MHz, CDCl3) δ 19.3, 27.3, 27.8, 31.4, 31.9, 32.1, 34.4, 43.2, 66.9, 77.0, 114.2, 117.4, 121.7, 126.4, 122.4, 137.1, 150.9, 152.6; MS (EI) m/z 300 (100), 298 (42), 285 (36), 207 (51); [αD20 −103° (c=10.4,CHCl3); HRMS calcd for C20H28O2: 300.2090, found 300.2089.
1-Deoxy-3-(1′,1′-dimethylbutyl)-11-hydroxy-Δ8-THC (9c). 11-Hydroxycannabinoid 9c was prepared by the procedure described above for the preparation of 9b. Stepwise oxidation and reduction of 1.20 g (4.09 mmol) of 8c 20 gave 0.33 g (25% for two steps)of hydroxy cannabinoid 9c as a white foam: 1H NMR (500 MHz, CDCl3) δ 0.81 (t, J=7.4 Hz,3H),1.03 .1.14 (m, 2H), 1.16 (s, 3H),1.25(s, 6H),1.40 (s, 3H), 1.49 .1.56 (m, 2H), 1.74 (td, J=5.0, 11.9 Hz, 1H), 1.82 .1.90 (m, 1H), 1.91 .2.02 (m, 2H), 2.23 (br.d, J=17.8 Hz,1H), 2.68 (td, J=5.5, 11.0 Hz,1H), 2.80 (dd, J=5.0, 17.0 Hz,1H), 4.04 (d, J=12.8 Hz,1H), 4.08 (d, J=12.8 Hz,1H), 5.75 (br.s,1H), 6.76 (d, J=1.8 Hz, 1H), 6.84 (dd, J=1.8, 8.2 Hz,1H),7.13 (d, J=7.8 Hz,1H); 3C NMR (75.5 MHz,CDCl3) δ 14.7, 17.9,19.1, 27.1, 27.6, 28.7, 28.8, 31.8, 32.0, 37.4, 43.0, 46.9, 66.8,7 6.6, 114.6, 117.7, 121.5, 122.0, 126.0, 136.9, 149.5, 152.4; MS (EI) m/z 328 (46), 285 (100), 207 (60); [αD20 −153°=10.3,CHCl3); HRMS calcd for C22H32O2: 328.2402, found 328.2402.
1-Deoxy-3-(1′,1′-dimethylpentyl)-11-hydroxy-Δ8-THC (9d). Hydroxycannabinoid 9d was prepared by the procedure described above for the preparation of 9b. Stepwise oxidation and reduction of 1.60 g (4.90 mmol)of 8d 20 gave 0.48 g (28% for two steps) of hydroxy cannabinoid 9d as a white foam: 1H NMR (500 MHz,CDCl3) δ 0.81 (t, J=7.3 Hz, 3H), 1.01 .1.09 (m, 2H), 1.17 (s, 3H), 1.17.1.23 (m,2H), 1.24 (s, 6H), 1.41 (s, 3H), 1.51 .1.57 (m, 2H), 1.76 (td, J=4.6, 11.4 Hz, 1H), 1.82.1.91 (m, 1H), 1.95 .2.03 (m, 2H), 2.25 (dt, J=8, 17.9 Hz, 1H), 2.69 (dt, J=5.5, 11.4 Hz, 1H), 2.80 (dd, J=4.6, 16.5 Hz, 1H), 4.06 (d, J=12.8 Hz,1H), 4.09 (d, J=12.8 Hz, 1H), 5.76 (br.s, 1H), 6.76 (d, J=1.8 Hz,1H), 6.84 (dd, J=1.8, 7.8 Hz,1H),7.14 (d, J=8.2 Hz, 1H); 13C NMR (125.8 MHz,CDCl3) δ 14.2, 19.2, 23.5, 27.0, 27.3, 27.8, 29.0, 31.9, 32.1, 37.4,43.1,44.4,67.1, 76.9, 114.8, 117.9, 121.8, 122.2, 126.2, 137.1, 149.8, 152.6; MS (EI) m/z 342 (33), 285 (100,), 269 (29), 255 (19); [αD20 +63° (c=14.3, CHCl3); HRMS calcd for C23H34O2: 342.2562, found 342.2559.
1-Deoxy-3-(1′,1′-dimethylhexyl)-11-hydroxy-Δ8-THC (9e). 11-Hydroxycannabinoid 9e was prepared by the procedure described above for the preparation of 9b. Stepwise oxidation and reduction of 1.72 g (5.05 mmol) of 8e21gave 0.231 g (13% for two steps)of hydroxy cannabinoid 9e as a white foam: 1H NMR (500 MHz, CDCl3) δ 0.82(t, J=6.8 Hz, 3H), 1.03 .1.12 (m,2H), 1.15 .1.24 (m, 4H), 1.16 (s, 3H), 1.24 (s, 6H), 1.40 (s, 3H), 1.51 .1.57 (m, 2H), 1.76 (td, J=4.6, 11.9 Hz,1H), 1.81 .2.03 (m, 3H), 2.23 (dt, J=8.0, 16.5 Hz, 1H), 2.69 (td, J=5.5, 11.0 Hz, 1H), 2.79 (dd, J=5.0, 17.0 Hz, 1H), 4.04 (d, J=12.8 Hz, 1H), 4.08 (d, J=12.8 Hz, 1H), 5.75 (br.s, 1H), 6.76 (d, J=2.3 Hz, 1H), 6.84 (dd, J=1.8, 8.2 Hz,1H),7.13 (d, J=8.2 Hz,1H); 13C NMR (125.8 MHz,CDCl3) δ 14.2, 19.2, 22.720 , 24.5, 27.3, 27.8, 28.9, 29.0, 31.9, 32.1, 32.7, 37.5, 43.2, 44.6, 67.0, 77.2, 114.8, 117.9, 121.7, 122.2, 126.2, 137.1, 149.8, 152.6; MS (EI) m/z 356 (32), 285 (100), 269 (18); [αD20 144 (c=6.5,CHCl3); HRMS calcd for C24H36O2: 356.2715, found 356.2715.
3-(1′,1′-Dimethylpropyl)-11-pivaloyloxy-Δ8-THC (13b). To a solution of 0.507 g (2.81 mmol) of crude 2-methyl-2-(3,5-dihydroxyphenyl) propane and 0.709 g (2.81 mmol) of 4-hydroxymyrtenyl pivalate (12) in 188 mL of dry dichloromethane at −20° C. was added dropwise with stirring 1.94 mL (14.1 mmol) of boron trifluoride etherate. The mixture was allowed to warm to 0° C. and stirred for 2 h, poured onto ice and neutralized with saturated aqueous NaHCO3. After extraction with ether, the combined organic extracts were dried (MgSO4) and concentrated in vacuo to afford the crude product as a dark brown foam. Chromatography (petroleum ether:ethyl acetate, 95:5) and subsequent recrystallization (heptane:ethyl acetate, 95:5)afforded 0.233 g (20%)of pure 13b as white crystals, mp 214-215° C.: 1H NMR (300 MHz,CDCl3) δ 0.67 (t, J=7.4 Hz, 3H), 1.10 (s, 3H),1.18 (s,6H),1.20 (s,9H),1.38 (s,3H),1.56 (q, J=7.3 Hz,2H), 1.80.1.91 (m, 3H), 2.21 .2.29 (m, 1H), 2.71 (td, J=4.6, 11.1 Hz, 1H), 3.36 (dd, J=3.9, 16.8 Hz, 1H), 4.48 (s, 2H), 5.20 (br.s, 1H), 5.73 (d, J=4.9 Hz, 1H),6.21 (d, J=1.8 Hz, 1H), 6.37 (d, J=1.7 Hz, 1H); 13C NMR (75.5 MHz,CDCl3) δ 9.2, 18.4, 27.2, 28.2, 31.2, 31.6, 36.8, 38.9, 44.8, 68.1, 76.5, 105.6, 107.8, 109.7, 123.2, 133.9, 149.7, 154.3, 154.7, 178.8; anal. calcd for C26H38O4: C,75.33; H,9.24; found:C,75.15; H,9.34.
1-Methoxy-3-(1′,1′-dimethylpropyl)-11-pivaloyloxy-Δ8-THC. Methylation of 0.556 g (1.34 mmol)of 13b by the procedure used for the preparation of 6b gave 0.454 g (79%) of the corresponding methyl ether as a colorless oil following chromatography (petroleum ether:ether, 97.5:2.5 to 95:5): 1H NMR (300 MHz,CDCl3) δ 0.70 (t, J=7.4 Hz, 3H), 1.10 (s, 3H), 1.24 (s, 15H), 1.39 (s, 3H), 1.57 (q, J=7.7 Hz, 2H),1.70 .1.92 (m, 3H), 2.21 .2.29 (m,1H), 2.67 (td, J=4.6, 11.1 Hz,1H), 3.30 (dd, J=3.9, 16.8 Hz,1H), 3.79 (s,3H), 4.49 (s, 2H), 5.76 (br.s,1H), 6.38 (d, J=1.5 Hz,1H), 6.43 (d, J=1.6 Hz,1H).
1-Methoxy-3-(1′,1′-dimethylpropyl)-11-hydroxy-Δ8-THC (11b). To a solution of 0.454 g (1.06 mmol)of pivalate ester in 33 mL of dry THF under N2 at 0° C. was added 0.051 g (1.34 mmol)of LiAlH4.The mixture was allowed to warn to room temperature, stirred for 1 h and quenched with 15 mL of aqueous NH4Cl. The solids were filtered off through a pad of Celite, which was subsequently washed with ether. The combined organic fractions were dried (MgSO4) and concentrated in vacuo to afford the crude product. Chromatography (gradient elution, petroleum ether:acetone, 95:5 to 93:7) gave 0.233 g (68%)of pure 11b as a colorless resin: 1H NMR (500 MHz,CDCl3) δ 0.69 (t, J=7.3 Hz, 3H), 1.11 (s,3H), 1.25 (s,6H), 1.39 (s, 3H), 1.59 (q, J=7.4 Hz, 2H),1.78 .1.91 (m, 3H), 2.18 .2.26 (m, 1H), 2.67 (td, J=4.6, 11.0 Hz,1H), 3.31 (dd, J=4.6, 16.5 Hz, 1H), 3.80 (s, 3H), 4.03 (d, J=13.3 Hz, 1H), 4.06 (d, J=13.3 Hz, 1H), 5.73 (d, J=4.6 Hz, 1H), 6.38 (d, J=1.6 Hz, 1H), 6.43 (d, J=1.6 Hz, 1H); 13C NMR (125.8 MHz,CDCl3) δ 9.3, 18.5, 27.6, 27.7, 28.4, 31.6, 31.9, 36.9, 38.0, 45.3, 55.2, 67.2, 76.6, 100.8, 108.4, 111.3, 120.8, 138.6, 149.5, 154.0,158.7, 158.7; MS (EI) m/z 344(29), 315(24), 281 (28), 245 (64), 207 (100); [αD20 −209° (c=0.46,CH2Cl2); HRMS calcd for C22H32O3 344.2351, found 344.2351.
3-(1′,1′-Dimethylethyl)-11-pivaloyloxy-Δ8-THC (13a). 11-Pivaloyloxycannabinoid 13a was prepared by the procedure described above for the preparation of 13b. From 1.62 g (9.76 mmol) of resorcinol 0.892 g (23%)of 13a was obtained as a colorless oil following chromatography (petroleum ether/ether, 97.5:2.5 to 95:5): 1H NMR (500 MHz,CDCl3) δ 1.05 (s, 3H), 1.15 (s, 9H), 1.16 (s, 9H), 1.32 (s, 3H), 1.73 .1.82 (m, 3H), 2.14.2.20 (m,1H), 2.63 (td, J=4.6, 11.0 Hz, 1H),3.30 (dd, J=4.2, 11.9 Hz, 1H), 4.43 (br.s, 2H), 5.35 (br. s, 1H), 5.68 (d, J=4.6 Hz, 1H), 6.24 (d, J=1.8 Hz, 1H), 6.36 (d, J=1.8 Hz, 1H); 13C NMR (125.8 MHz,CDCl3) δ 18.6, 27.3, 27.6, 27.7, 31.3, 31.4, 34.4, 39.0, 44.8, 68.3, 76.9, 105.1, 107.2, 109.9, 123.3, 133.9, 151.4, 154.5, 154.9, 178.9, 154.9.
1-Methoxy-3-(1′,1′-dimethylethyl)-11-pivaloyloxy-Δ8-THC. Methylation of 0.523 g (1.31 mmol)of 13a by the procedure used for the preparation of 6b gave 0.447 g (82%) of the corresponding methyl ether as a colorless oil following chromatography (petroleum ether/ether, 97.5:2.5 to 95:5): 1H NMR (300 MHz,CDCl3) δ 1.11 (s, 3H), 1.23 (s, 9H), 1.29 (s, 9H), 1.39 (s, 3H), 1.74.1.87 (m, 3H), 2.19.2.30 (m,1H), 2.67 (td, J=4.5, 10.9 Hz, 1H), 3.31 (dd, J=3,9,16.9 Hz, 1H), 3.80 (s,3H), 4.49 (br s,2H), 5.75 (d, J=4.4 Hz, 1H), 6.44 (d, J=1.6 Hz, 1H), 6.49 (d, J=1.6 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 18.4, 27.2, 27.5, 27.7, 31.2, 31.4, 31.8, 39.0, 44.9, 55.0, 68.1, 76.3, 100.2, 107.6, 111.2, 123.4, 134.1, 149.9, 154.0, 158.7, 178.4.
1-Methoxy-3-(1′,1′-dimethylethyl)-11-hydroxy-Δ8-THC (11a). Reduction of 0.447 g (1.08 mmol)of the pivalate ester by the procedure used for the preparation of 11b gave 0.176 g (49%) of hydroxy cannabinoid 11a as a colorless resin: 1H NMR (500 MHz,CDCl3) δ 1.10 (s, 3H), 1.27 (s, 9H), 1.38 (s, 3H), 1.76.1.90 (m, 3H), 2.17.2.24 (m,1H), 2.65 (td, J=4.6, 11.0 Hz,1H), 3.30 (dd, J=4.6, 16.5 Hz, 1H), 3.80 (s, 3H),4.02(d, J=13.3 Hz,1H),4.05 (d, J=13.3 Hz,1H), 5.72(d, J=5.0 Hz,1H), 6.43 (d, J=1.9 Hz, 1H), 6.48 (d, J=1.8 Hz, 1H); 13C NMR (125.8 MHz,CDCl3) δ 18.5, 27.7, 31.3, 31.5, 31.8, 34.8, 45.2, 55.2, 67.2, 76.6, 100.2, 107.6, 111.4, 120.8, 138.6, 151.1, 154.0, 158.8; MS (EI) m/z 330 (34), 299 (14), 231 (100), 207 (92); [αD20 −225° (c=1.08,CH2Cl2); HRMS calcd for C21H30O3: 330.2194, found 330.2195.
3-(1′,1′-Dimethylbutyl)-1-methoxy-11-oxo-Δ8-THC (14c). Selenium dioxide oxidation of 0.290 g (0.847 mmol)of 6c, using the procedure described above for the preparation of 10b gave, after chromatography (petroleum ether/ethyl acetate, 9:1), 0.196 g (65%)of aldehyde as a yellow solid:mp 109-110° C.; 1H NMR (300 MHz,CDCl3) δ 0.82(t, J=7.2 Hz,3H),1.03 .1.13 (m,2H), 1.10 (s,3H),1.25 (s,6H),1.43 (s,3H),1.50 .1.55 (m, 2H),1.79 .1.93 (m.2),2.04 .2.15 (m,1H),2.51 .2.56 (m, 1H),2.61 (td, J=4.5, 11.2 Hz,1H),3.73 (dd, J=2.1, 17.9 Hz, 1H), 3.82, (s,3H), 6.39 (d, J=1.6 Hz,1H),6.42 (d, J=1.6 Hz,1H), 6.83 (s,1H), 9.50 (s,1H); 3C NMR (75.5 MHz,CDCl3) δ 14.8, 17.9, 18.2, 27.5, 28.7, 29.2, 30.8, 37.6, 44.9, 47.0, 55.1, 75.8, 100.7, 108.2, 110.2, 142.2, 148.9, 150.1, 153.7, 158.6, 193.7; MS (EI) m/z 356 (55), 306 (100).
1-Methoxy-3-(1′,1′-dimethylbutyl)-11-hydroxy-Δ8-THC (11c). To a suspension of 0.196 g (0.551 mmol)of aldehyde 14c in 3.5 mL of dry methanol was added sequentially 0.205 g (0.551 mmol)of CeCl3.7H2O and 0.021 g (0.551 mmol)of NaBH4.The reaction mixture was stirred at ambient temperature for 2 h, the pH was adjusted to 7.0 by the addition of 1M aqueous HCl. After pouring into water the mixture was extracted with three portions of CH2Cl2,dried (MgSO4)and the solvent was removed in vacuo. The residue was purified by chromatography (petroleum ether:ethyl acetate, 4:1) to give 0.185 g (94%) of cannabinoid 11c as a pale yellow gum: 1H NMR (500 MHz,CDCl3) δ 0.82 (t, J=7.1 Hz,3H), 1.05.1.14 (m,2H),1.11 (s,3H),1.25 (s,6H),1.39 (s, 3H),1.50 .1.54 (m,2H),1.79 .1.92 (m,3H),2.17 .2.24 (m,1H),2.67 (td, J=4.6, 11.0 Hz,1H),3.31 (dd, J=4.6, 16.5 Hz,1H),3.80 (s,3H),4.05 (s,2H),5.73 (s,1H), 6.38 (d,1.6.1H),6.43 (d, J=1.6 Hz,1H); 13C NMR (125.8 MHz,CDCl3) δ 14.7,17.9,18.3,27.5,27.6,28.7, 28.8, 31.4, 31.8, 37.6, 45.1, 46.9, 55.0, 66.8, 76.2, 100.5, 108.1,111.1,120.4,138.4,149.6,153.8,158.5; [αD20 −238° (c=0.50,CHCl3); HRMS calcd for C23H34O3:358.2508, found 358.2509.
1-Methoxy-3-(1′,1′-dimethylpentyl)-11-oxo-Δ8-THC (14d). Selenium dioxide oxidation of 2.00 g (5.17 mmol) of 6d, using the procedure described above for the preparation of 10b, gave, after chromatography (petroleum ether/ethyl acetate, 87.5:12.5), 1.17 g (50%) of aldehyde 14d as a pale orange solid: 1H NMR (300 MHz,CDCl3) δ 0.83 (t, J=7.4 Hz,3H),1.02.1.09 (m,2H),1.14 (s, 3H),1.18.1.28 (m,2H),1.25 (s,6H),1.43 (s,3H),1.52.1.60 (m,2H), 1.76.1.94 (m,2H),2.05.2.19 (m,1H), 2.53.2.58 (m,1H),2.61 (td, J=4.5, 11.2 Hz,1H),3.74 (dd, J=3.6, 16.1 Hz,1H),3.83 (s,3H),6.39 (d, J=1.6 Hz,1H),6.43 (d, J=1.6 Hz,1H),6.83.6.84 (m, 1H),9.50 (s,1H); 13C NMR (75 MHz,CDCl3) 514.1, 18.2,23.4,26.9,27.6,28.7,28.8,29.2,30.8,37.9,44.2, 45.0,47.3,55.1,75.8, 100.7, 108.1, 110.2, 142.5, 148.9, 150.2,153.7,158.6,193.8; MS (EI)m/z 370 (35),314 (100), 300 (25).
1-Methoxy-3-(1′,1′-dimethylpentyl)-11-hydroxy-Δ8-THC (11d). Luche reduction of 0.500 g (1.35 mmol)of aldehyde 14d by the procedure described above for the reduction of 14c gave after chromatography (petroleum ether/ethyl acetate,4:1) 0.350 g (70%)of 11-hydroxy cannabinoid 11d as a pale yellow gum: 1H NMR (300 MHz,CDCl3) δ 0.83 (t, J=7.3 Hz,3H),1.00.1.10 (m,2H), 1.11 (s,3H),1.16.1.28 (m,2H),1.24 (s,6H), 1.39 (s,3H),1.49.1.57 (m,2H),1.77.1.91 (m,3H), 2.20.2.25 (m,1H),2.66 (td, J=4.6, 10.9 Hz,1H),3.31 (dd, J=3.6, 17.1 Hz,1H),3.80 (s,3H),4.05 (s,2H),5.73 (d, J=4.2 Hz,1H),6.37 (d, J=1.6 Hz,1H),6.42(d, J=1.6 Hz,1H); 13C NMR (75.5 MHz,CDCl3) δ 14.1, 18.4,23.4, 26.9, 27.6, 27.7, 28.8, 31.5, 31.8,37.6, 44.2,45.2,67.1, 76.3, 100.6, 108.2, 111.1, 120.7, 138.5, 149.8, 153.8,158.6;[αD20 −284° (c=0.25, CHCl3); HRMS calcd for C24H36O3: 372.2664, found 372.2663.
1-Methoxy-3-(1′,1′-dimethylhexyl)-11-oxo-Δ8-THC (14e). Selenium dioxide oxidation of 2.25 g (6.07 mmol) of 6e, using the procedure described above for the preparation of 10b gave 1.40 g (60%)of aldehyde 14e as an orange solid, mp 117-119° C.; 1H NMR (300 MHz, CDCl3) δ 0.83 (t, J=7.1 Hz,3H),1.02.1.30 (m,6H), 1.10 (s,3H),1.25 (s,6H),1.43 (s,3H),1.51.1.56 (m, 2H),1.80.1.93 (m,2H),2.05.2.18 (m,1H),2.49.2.56 (m,1H),2.62 (td, J=4.5, 11.2 Hz,1H),3.73 (dd, J=4.1, 15.7 Hz,1H),3.82(s,3H),6.39 (d, J=1.6 Hz,1H),6.42 (d, J=1.6 Hz,1H),6.84 (br.s,1H), 9.50 (s,1H);13C NMR (75.5 MHz,CDCl3) δ 14.1,18.2, 22.5, 24.3, 27.5, 28.7, 28.8, 29.3, 30.9, 32.5, 37.7, 44.5, 44.9, 55.2,75.8, 100.7, 108.1, 110.3, 142.5, 148.9, 150.2, 153.7, 158.7, 193.8; MS (EI) m/z 384 (20), 314 (100).
1-Methoxy-3-(1′,1′-dimethylhexyl)-11-hydroxy-Δ8-THC (11e). Luche reduction of 0.404 g (1.05 mmol)of aldehyde 14e by the procedure described above for the reduction of 14c gave after chromatography (petroleum ether/ethyl acetate, 4:1) 0.354 g (87%)of 11-hydroxy cannabinoid 11e as a pale yellow oil: 1H NMR (300 MHz,CDCl3) δ 0.83 (t, J=7.3 Hz,3H),1.07 .1.34 (m,6H),1.11 (s,3H), 1.25 (s,6H), 1.40 (s,3H), 1.51.1.58 (m,2H),1.78.1.88 (m,3H),2.21 .2.25 (m,1H),2.67 (td, J=4.6, 10.9 Hz,1H),3.31 (dd, J=3.5, 17.2 Hz,1H), 3.81 (s,3H), 4.06 (s,2H),5.73 (d, J=4.4 Hz,1H),6.38 (d, J=1.6 Hz,1H),6.43 (d, J=1.6 Hz,1H); 13C NMR (75.5 MHz,CDCl3) δ 14.1, 18.4, 22.6, 24.3, 27.6, 27.7, 28.8, 28.9, 31.5, 31.9, 32.6, 44.4, 45.2, 55.2, 67.2,76.3, 100.7, 108.3, 111.2, 120.8, 138.6, 149.9, 154.0, 158.7; HRMS calcd for C25H38O3 386.2821, found 386.2818.
3-(1′,1′-Dimethylheptyl)-1-methoxy-11-oxo-Δ8-THC (14f). Selenium dioxide oxidation of 1.64 g (4.27 mmol) of 4, using the procedure described above for the preparation of 10b gave 1.12g (66%) of aldehyde 14f as an orange solid: mp 118-119° C.; Rf 0.31 (petroleum ether/ethyl acetate, 94:6); 1H NMR (300 MHz,CDCl3) δ 0.84 (t, J=6.9 Hz,3H),1.06.1.27 (m,8H),1.14 (s,3H),1.24 (s,6H),1.43 (s,3H),1.51.1.57 (m,2H),1.66.1.68 (m, 1H),1.88 (td, J=4.1, 11.6 Hz,1H),2.07.2.18 (m,1H), 2.51.2.57 (m,1H),2.61 (td, J=4.1, 11.2 Hz,1H),3.73 (dd, J=4.5, 17.1 Hz,1H),3.81 (s,3H),6.39 (d, J=1.6 Hz,1H),6.42(d, J=1.6 Hz,1H),6.83 (s,1H), 9.50 (s,1H);13C NMR (75.5 MHz,CDCl3) δ 14.1, 18.2, 22.6, 24.6, 27.5, 28.7, 28.9, 29.2, 30.0, 30.8, 31.7, 37.7, 44.5, 45.0, 55.1, 75.8, 100.7, 108.1, 110.2, 142.5, 148.9, 150.2, 153.7, 158.6, 193.8; MS (EI)m/z 398 (25), 314 (50), 281 (66), 207 (100).
11-Hydroxy-3-(1′,1′-dimethylheptyl)-1-methoxy-D 8-THC (11f).Luche reduction of 0.314 g (0.789 mmol)of aldehyde 14f by the procedure described above for the preparation of 14c gave after chromatography (petroleum ether/ethyl acetate, 85:15) 0.270 g (93%)of 11-hydroxy cannabinoid 11f as a pale yellow oil: Rf 0.30 (petroleum ether/ethyl acetate,85:15); 1H NMR (300 MHz,CDCl3) δ 0.84 (t, J=6.9 Hz, 3H), 0.95.1.26 (m, 8H), 1.10 (s, 3H), 1.24 (s, 6H), 1.39 (s, 3H), 1.51.1.56 (m, 2H), 1.78.1.90 (m, 4H), 2.19.2.26 (m, 1H), 2.66 (td, J=4.1, 10.8 Hz,1H), 3.30 (dd, J=3.3, 16.6 Hz,1H), 3.79 (s,3H), 4.03 (br s,2H), 5.71 (s,1H), 6.37 (d, J=1.6 Hz,1H), 6.42(d, J=1.6 Hz,1H); 13C NMR (75.5 MHz,CDCl3) δ 14.0,18.3, 22.6,24.5, 27.5,27.6, 28.7, 28.8, 29.6, 29.9, 31.4, 31.7, 37.6, 44.4, 45.1, 55.0, 66.9, 76.3, 100.6, 108.2, 111.1, 120.5, 138.5, 149.7, 153.8, 158.6; [αD20 −175° (c=0.3,CH2Cl2); HRMS calcd for C26H4O3: 400.2977, found 400.2979.
Receptor Binding Assays
1.CB1 assay.[3H] CP-55,940 (KD32 690 nM) binding to P2 membranes was conducted as described elsewhere,31 except whole brain (rather than cortex only) was used. Displacement curves were generated by incubating drugs with 1 nM of [3H]CP-55,940. The assays were performed in triplicate, and the results represent the combined data from three individual experiments.
2.CB2assay. Human embryonic kidney 293 cells were maintained in Dulbecco's modified Eagle's medium (DMEM) with 10% fetal clone II (HyClone, Logan Utah)and 5% CO2 at 37° C. in a Forma incubator. Cell lines were created by transfection of CB2 pcDNA3into 293 cells by the Lipofectamine reagent (Life Technologies, Gaithersburg, Md.).The human CB2 cDNA was a gift. Stable transformants were selected in growth medium containing geneticin (1 mg/mL, reagent, Life Technologies, Gaithersburg, Md.). Colonies of about 500 cells were picked (about 2 weeks post transfection) and allowed to expand, then tested for expression of receptor mRNA by northern blot analysis. Cell lines containing moderate to high levels of receptor mRNA were tested for receptor binding properties. Transfected cell lines were maintained in DMEM with 10% fetal clone II plus 0.3-.0.5 mg/mL geneticin and 5% CO2 at 37° C. in a Forma incubator.
The current assay is a modification of Compton et al.8 Cells were harvested in phosphate-buffered saline containing 1 mM EDTA and centrifuged at 500 g. The cell pellet was homogenized in 10 mL of solution A (50 mM Tris-HCl, 320 mM sucrose, 2 mM EDTA, 5 mM MgCl2, pH 7.4). The homogenate was centrifuged at 1600 g (10 min),the supernatant saved, and the pellet washed three times in solution A with subsequent centrifugation. The combined supernatants were centrifuged at 100,000 g (60 min). The (P2 membrane) pellet was resuspended in 3 mL of buffer B (50 mM Tris-HCl, 1 mM EDTA, 3 mM MgCl2, pH 7.4) to yield a protein concentration of approximately 1 mg/mL. The tissue preparation was divided into equal aliquots, frozen on dry ice, and stored at 70° C. Binding was initiated by the addition of 40-50 μg membrane protein to silanized tubes containing [3H] CP-55,940 (102.9 Ci/mmol) and a suffcient volume of buffer C (50 mM Tris-HCl,1 mM EDTA,3 mM MgCl2, and 5 mg/mL fatty acid free BSA, pH 7.4) to bring the total volume to 0.5 mL. The addition of 1 m unlabelled CP-55,940 was used to assess nonspecific binding. Following incubation (30° C. for 1 h),binding was terminated by the addition of 2 mL of ice cold buffer D (50 mM Tris-HCl, pH 7.4, plus 1 mg/mL BSA) and rapid vacuum filtration through Whatman GF/C filters (pretreated with polyethyleneimine (0.1%) for at least 2 h).Tubes were rinsed with 2 mL of ice cold buffer D, which was also filtered, and the filters subsequently rinsed twice with 4 mL of ice cold buffer D. Before radioactivity was quantitated by liquid scintillation spectrometry, filters were shaken for 1 h in 5 mL of scintillation fluid.
CP-55,940 and all cannabinoid analogues were prepared by suspension in assay buffer from a 1 mg/mL ethanolic stock without evaporation of the ethanol (final concentration of no more than 0.4%). When anandamide was used as a displacing ligand, experiments were performed in the presence of phenylmethylsulfonyl fluoride (50 μM). Competition assays were conducted with 1 nM [3H] CP-55,940 or 1 nM [3H] SR141716A and 6 concentrations (0.1 nM to 10 m displacing ligands). Displacement IC50values were originally determined by unweighted least-squares linear regression of log concentration-percent displacement data and then converted to Ki values using the method of Cheng and Prusoff.32
Abstract: The synthesis, CB1 and CB2 receptor affinities of 47 cannabimimetic indoles are described. The indole derivatives include 1-propyl- and 1-pentyl-3-(1-naphthoyl)indoles both with and without a 2-methyl substituent. Naphthoyl substituents include 4- and 7-alkyl groups as well as 2-, 4-, 6-, 7-methoxy and 4-ethoxy groups. The effects of these substituents on receptor affinities are discussed and structure-activity relationships are presented. In the course of this work three new highly selective ligands for the CB2 receptor were identified, 1-propyl-3-(4-methyl-1-naphthoylindole (JWH-120), 1-propyl-2-methyl-3-(6-methoxy-1-naphthoylindole (JWH-151), and 1-pentyl-3-(2-methoxy-1-naphthoylindole (JWH-267).
Introduction
Nearly 40 years ago Gaoni and Mechoulam reported the elucidation of the structure of Δ9-tetrahydrocannabinol (1, THC, Scheme 1) the principal psychoactive compound present in marijuana.1 Subsequently, a comprehensive set of structure-activity relationships (SAR) were developed based on the dibenzopyran nucleus of THC.2-6 These SAR were later extended to the very potent group of non-traditional cannabinoids developed by Pfizer, of which CP-55,940 (2, DMH=1,1-dimethylheptyl, Scheme 1) is the prototypical example.7,8
These pharmacological effects of cannabinoids are considered to be mediated through at least two G-protein-coupled, transmembrane receptors. One of these, designated as CB1, is found predominantly in the central nervous system and is thought to be responsible for most of the overt pharmacological effects of cannabinoids.5,9-11 A second receptor, designated CB2 was originally identified from macrophages present in the spleen, and is expressed primarily in the periphery.12 Very recently evidence has been presented for the existence of a third cannabinoid receptor, which has been detected in mouse brain.13
Some years ago in the course of a program directed toward the development of nonsteroidal anti-inflammatory drugs, a group at Sterling-Winthrop reported that pravadoline (3), an indole derivative, unexpectedly inhibited contractions of the electrically stimulated mouse vas deferens.14 Later work revealed that 3 (Scheme 1), and a number of related compounds also inhibit adenylate cyclase, are antinociceptive and interact with a G-coupled protein in the brain. Subsequent work indicated that the G-coupled protein is the cannabinoid CB1 receptor and that these aminoalkylindoles exhibit typical cannabinoid pharmacology in vivo.15,16 One of the aminoalkylindoles, WIN-55,212-2 (4, Scheme 1), developed by the Winthrop group is not only potent in vivo, but has high affinity for both the cannabinoid CB1 and CB2 receptors.17
During the course of their investigations of the aminoalkylindoles, the Winthrop group synthesized more than 100 compounds and developed some preliminary SAR.14,15,18 These included the observation that a group larger than methyl at C-2 of the indole nucleus greatly attenuates potency and that a bicyclic aroyl group, usually 1-naphthoyl or a substituted 1-naphthoyl group, at C-3 is essential for potency. It was further concluded that an aminoalkyl group, usually substituted aminoethyl, attached to the indole nitrogen was essential for cannabinoid activity.
Subsequent studies by our group established that the aminoalkyl group appended to the indole nitrogen could be replaced by an alkyl group to provide relatively simple indole derivatives which exhibit typical cannabinoid pharmacology.19,20 In particular, JWH-007, 1-pentyl-2-methyl-3-(1-naphthoyl)indole (5, Scheme 1) has high affinity for the CB1 receptor and exhibits typical cannabinoid pharmacology in vivo. The 1-propyl analog of 5, JWH-015 (6, Scheme 1) has relatively high affinity for the CB2 receptor, and weak affinity for the CB1 receptor.17
In an effort to develop improved indole based CB2 cannabinoid receptor ligands and also to develop SAR for both the CB1 and CB2 receptors a number of additional indole derivatives were prepared and their pharmacology was evaluated.21,22 It was found that CB1 receptor affinity is optimal with a n-pentyl nitrogen substituent, and decreases dramatically with N-alkyl substituents of three or less carbon atoms. CB1 receptor affinity is also greatly attenuated with N-alkyl substituents longer than six carbon atoms. A 2-methyl substituent slightly decreases affinity at the CB1 receptor and a 4-methoxy-1-naphthoyl group at C-3 of the indole, as in JWH-098 (7) enhances CB1 receptor affinity. A 7-methyl-1-naphthoyl substituent in JWH-046 (8) and JWH-048 (9) has relatively little effect on either CB1 or CB2 receptor affinity. The SAR at the CB2 receptor are somewhat similar to those at the CB1 receptor, however there are a number of exceptions to these generalizations which render it difficult to establish comprehensive SAR for these compounds at the CB2 receptor.21,23
The SAR outlined above includes only indoles with an unsubstituted 3-naphthoyl substituent, or with 4-methoxy- or 7-methyl-1-naphthoyl groups. To further develop SAR for these compounds, and with the goal of preparing CB2 selective ligands, a number of additional cannabimimetic indoles have been prepared and their affinities for the CB1 and CB2 receptors have been determined. These compounds include N-propyl- and N-pentyl-3-(1-naphthoyl) indoles in which the naphthoyl group contains various alkyl and alkoxy substitutuents, and the indole is either unsubstitued at C-2 or contains a 2-methyl group. The choice of the N-propyl substituent is based on the observation that the two most highly CB2 selective indole derivatives prepared in our laboratory, JWH-015 (6) and JWH-046 (8) both contain this substitution pattern.21,23 The N-pentyl group was chosen since this substituent almost invariably provides compounds with higher CB1 receptor affinities than are observed with other nitrogen substituents.
Results
To explore the steric and electronic effects of various naphthoyl substituents, 3-(4-alkyl-1-naphthoyl), 3-(7-ethyl-1-naphthoyl), 3-(2-, 6-,7-methoxy-1-naphthoyl), 3-(4-ethoxy-1-naphthoyl)indoles (for numbering see the structure included in Table 2), have been synthesized and their CB1 and CB2 receptor affinities have been determined. These indoles were prepared by modifications of established routes, either from substituted 1-naphthoyl chlorides and 1-alkyl- or 1-alkyl-2-methylindoles or by acylation of indole or 2-methylindole, followed by N-alkylation (Scheme 1).14,15,18,19,24
aRef 41.
bRef 17.
cRefs. 20 and 21.
dRef. 19.
eRef. 24.
fRef. 21.
In our early work, indole (10, R═H) or 2-methylindole (10, R═CH3) was treated with methylmagnesium bromide to give the ambident indolyl anion which upon reaction with an aroyl chloride gave the 3-acylindole (11, method A).19 N-Alkylation with the appropriate primary alkyl bromide using KOH in DMSO provides the N-alkyl-3-aroylindole. Although this sequence was satisfactory when using readily available aroyl halides, the yields in the first step were variable, the 3-aroylindoles (11) were difficult to purify and alternative methodology was explored. Traditional aluminum chloride catalyzed Friedel-Crafts acylation of N-alkylindoles gave poor yields, however two variations of this classical reaction gave satisfactory to good yields. The method of choice is that reported recently by Okauchi et al. (method B) in which N-alkylindoles 12 are treated with dimethyl- or diethylaluminum chloride for 30 minutes prior to the addition of the aroyl halide.25 This mild procedure provides pure products (13) in yields of 50% to 90%. The final methods are variations of a classical Friedel-Crafts procedure, but use ethylaluminum dichloride at ambient temperature for either four days (method C) or for 18 h (method D). The yields for procedures C and D are adequate, but are somewhat inferior to those using Okauchi's method (procedure B).
Of the 1-naphthoic acids required for the synthesis of the cannabimimetic 3-(4-alkyl-1-naphthoyl)indoles, only 4-methyl-1-naphthoic acid (14, Scheme 2) is commercially available. 4-Ethyl-(15), 4-propyl-(16) and 4-butyl-1-naphthoic (17) acids were reported some years ago by Russian workers, who prepared them by chloromethylation of the 1-alkylnaphthalene, followed by conversion to the corresponding acetate, hydrolysis to the alcohol and oxidation to the carboxylic acid.26 A considerably more efficient approach employs Friedel-Crafts acylation of the appropriate 1-alkylnaphthalene (18, R═C2H5, C3H7, C4H9) with N,N,-diphenylcarbamyl chloride to give 1-naphthyl N,N,-diphenylamides (19, R═C2H, C3H7, C4H9), hydrolysis of which will afford the corresponding carboxylic acid.27 Although the acylation step proceeded smoothly in yields of 80% to 85%, hydrolysis could not be effected using the reported procedure, ethanolic sodium hydroxide at reflux. The amides were successfully hydrolyzed in 90% to 92% yield by treatment with potassium hydroxide in diethylene glycol at reflux.28,29 naphthoic acids (14-17). Conversion of 4-methyl-1-naphthoic acid to 1-propyl-3-(4-methyl-1-naphthoyl)indole (JWH-120) and 2-methyl-1-propyl-3-(4-methyl-1-naphthoyl)indole (JWH-148) was carried out using methods B and A, respectively. The 1-pentyl analogs of these compounds (JWH-122, JWH-149) have been reported previously.24 The conversion of acids 14-17 to the 3-(4-ethyl-1-naphthoyl)indoles (JWH-210-JWH-213), the 3-(4-propyl-1-naphthoyl)indoles (JWH-180-JWH-182, JWH-189) and the 3-(4-butyl-1-naphthoyl)indoles (JWH-239-JWH-242) was carried out using method C. The CB1 and CB2 receptor affinities of these compounds are summarized in Table 2.
7-Methyl-1-naphthoic acid (20), which had been used previously for the synthesis of JWH-046 (8) and JWH-048 (9),21 was prepared by a modification of the method of Snatzke and Kunde (Scheme 3).30 Friedel-Crafts acylation of 2-methylnaphthalene (21) under standard conditions using acetyl chloride and AlCl3 proceeds as described to provide 1-acetyl-7-methylnaphthalene (22), however, conversion to acid 20 could not be effected using the haloform reaction under various conditions. In the King modification of the haloform reaction, ketone 22 with iodine and pyridine gave pyridinium iodide 23, which upon basic hydrolysis and acidification provided acid 20.31 Conversion of this acid to 1-propyl-3-(7-methyl-1-naphthoyl)indole was carried out by method A. 7-Ethyl-1-naphthoic acid (24), which was employed for the synthesis of the 3-(7-ethyl-1-naphthoyl)indoles was prepared from 2-ethylnaphthalene (25) in an analogous manner through ketone 26 and pyridinium iodide 27. Conversion of 7-ethyl-1-naphthoic acids to the 3-(7-ethyl-1-naphthoyl)indoles (JWH-234-JWH-236, JWH-262) was carried out using method B. The receptor affinities for the 3-(7-alkyl-1-naphthoyl)indoles are included in Table 2.
In the methoxynaphthoyl series, the 2-, 4-, 6- and 7-methoxy-1-naphthoylindoles were prepared and their affinities for the CB1 and CB2 receptors were determined (Table 3). In addition, to evaluate steric and electronic effects upon receptor affinities, the 4-ethoxy analogs were also prepared. The 3-(4-ethoxy-1-naphthoyl)indoles were prepared from 4-ethoxy-1-naphthoic acid (28, Scheme 2), which was obtained from 1-ethoxynaphthalene by the procedure outlined in Scheme 2. Acid 28 was converted to the 3-(4-ethoxy-1-naphthoyl)indoles (JWH-258-JWH-261) by method B and their receptor affinities are included in Table 3. The 3-(4-methoxy-1-naphthoyl)indoles, JWH-079, JWH-094, JWH-081, JWH-098, have been reported previously and their affinities for the CB1 and CB2 receptors are also included in Table 2.21,24
aRef. 41.
bRef. 17.
cRef. 21.
dRef. 24
For the synthesis of the 3-(2-methoxy-1-naphthoyl)indoles 2-methoxy-1-naphthoic acid (29) was required. Acid 29 is readily available 2-hydroxy-1-naphthoic acid 32 and was converted to Indoles 30-33 (JWH-265-JWH-268, Table 3) by method B. Indoles 30 (JWH-265) and (JWH-267) which are unsubstituted at C-2 of the indole nucleus show normal 1H and 13C NMR spectra, however in the 1H spectra of the 2 -methyl analogs (31, JWH-266, and 33, JWH-268) the signal at d 2.50 due to the indole 2-methyl group and an aromatic proton at d 6.95 appeared as very broad singlets. A second aromatic proton at d 7.13 gave rise to a somewhat broadened singlet, rather than the sharp signals characteristic of the naphthoylindoles. Also, the signals due to the 2-methyl group and one aromatic carbon were missing in the 13C spectra of indoles 31 and 33. Although it was possible that these compounds did not have the correct structures, both had low and high resolution mass spectra consistent with the assigned molecular formulas. Ultimately the structure of 1-propyl-2-methyl-3-(2-methoxy-1-naphthoyl)indole (31) was established by x-ray crystallography.33
It appeared probable that the anomalous NMR spectra were due to restricted rotation about the bond between C-3 of the indole nucleus and/or the naphthalene C-1 bond and the carbonyl carbon atom. Accordingly, a variable temperature study of the 1H NMR spectrum of indole 31 was carried out. There was little change in the 1H spectrum at 50° C., but at 75° C. the signal for the C-2 methyl group had sharpened somewhat, as had that of the aromatic proton at d 6.95, and the signal at d 7.13 was resolved to a broadened triplet, J=8.2 Hz. At 125° C. the C-2 methyl signal was a rather sharp singlet at d 2.46, the aromatic proton at d 6.95 had become a well defined triplet, J=7.8 Hz, and the triplet at d 7.13 appeared as a triplet of doublets, J=0.5 and 8.2 Hz. The 1H spectrum at 100° C. was slightly less well resolved than that at 125° C.
A COSY spectrum of the aromatic region of indole 31 revealed that the protons which give rise to the triplets at d 6.95 and d 7.13 are coupled to each other and each is coupled to one additional proton within the complex pattern between d 7.3 and v 7.6. The triplets at d 6.95 and d 7.13 can only arise from the 5,6-protons on the indole nucleus or the 6,7-naphthalene protons. Since the aromatic protons of a simple acyl indole derivative, 2-methyl-1-morpholinoethyl-3-acetylindole (34) appear as a series of multiplets between d 7.26 and d 7.97,34 the triplets at d 6.95 and d 7.13 in indole 31 must arise from the protons at C-6 and C-7 of the naphthalene. These data confirm that the anomalies in the NMR spectra of indoles 31 and 33 are the result of restricted rotation about the bond between C-3 of the indole nucleus and/or the naphthalene C-1 bond and the carbonyl carbon atom. The energy barrier to rotation was determined using variable temperature 1H NMR, and the coalescence temperature was found to be 0° C. The free energy of activation was calculated from the stopped exchange limit at 31 50° C. and the coalescence temperature to give a rotational barrier of 13.1 kcal/mol.35 An AMI coordinate drive study shows that during rotation about the naphthalene C-1 bond and the carbonyl carbon atom the oxygen of the methoxy group has close Van der Waals' contacts with C-2 and C-3 of the indole. The maximum overlap occurs at a carbonyl oxygen-carbonyl carbon-indole C-3-indole C-2 torsion angle of −89.8°. The calculated energy difference between the global minimum energy conformer and this conformation is 9.38 kcal/mol. Single point energy calculations for the difference in energy between the global minimum energy conformer and the conformation for which maximum overlap occurs was found to be 12.86 kcal/mol at the HF 3-21G* level. The receptor affinities of the 3-(2-methoxy-1-naphthoyl)indoles (JWH-265-JWH-268) are included in Table 3.
The 6- and 7-methoxy-1-naphthoylindoles required 6-(35, Scheme 4) and 7-methoxy-1-naphthoic acid (36), respectively, both of which were prepared a number of years ago using classical chemistry. The 6-methoxy acid (35) was originally synthesized by Price et al. in 12% yield by the aluminum chloride catalyzed reaction of furoic acid and anisole.36 7-Methoxy-1-naphthoic acid (36) was described by Fieser and Holmes who prepared it from ethyl 4-(4-methoxyphenyl)butanoate via 7-methoxy-3,4-dihydro-1-naphthoic acid.37 The dihydro acid was synthesized in 51% overall yield, however no yield was specified for the dehydrogenation to acid 36. A subsequent synthesis of acid 36 utilized the reaction of 2-methoxynaphthalene with an oxalyl chloride equivalent to give a methoxyacenapthenquinone, which was converted to acid 36 in two steps and good yield.38 In our hands attempted repetition of the reaction of 2-methoxynaphthalene with the oxalyl chloride surrogate failed. Other published syntheses of both acids did not appear attractive and alternative methods of synthesis were explored.
Both 6-(37) and 7-methoxy-1-tetralone (38, Scheme 4) are commercially available, and appeared to be suitable starting materials for the synthesis of acids 35 and 36. The initial synthetic route to 6-methoxy-1-naphthoic acid (35) entailed conversion of tetralone 37 to the enol triflate, followed by palladium mediated carbonylation to 6-methoxy-3,4-dihydro-1-naphthoic acid.39 Dehydrogenation to acid 35 did not proceed to completion and the mixture of 35 and the dihydro acid proved exceedingly difficult to separate. The conversion of tetralones 37 and 38 to 6-(39) and 7-methoxy-1-naphthol (40), respectively was reported some years ago by Shand and Thompson who accomplished this transformation by dehydrogenation using sulfur at 250° C.40 As an alternative to thermal dehydrogenation, ketones 37 and 38 were converted to the corresponding a-bromo ketones, dehydrohalogenation of which provided phenols 39 and 40.41 Conversion of these phenols to the corresponding triflates followed by palladium mediated carbonylation gave acids 35 and 36 in good yield. Acid 35 was converted to the 3-(6 -methoxy-1-naphthoyl)indoles (JWH-151, JWH-153, JWH163, JWH-166) and acid 36 was converted to the 3-(7-methoxy-1-naphthoyl)indoles (JWH-159, JWH-160, JWH-164, JWH-165) by method D. The receptor affinities for these 3-(6- and 7-methoxy-1-naphthoyl)indoles are included in Table 3.
The affinities of the alkylnaphthoyl and alkoxynaphthoylindoles for the CB1 receptor were determined by measuring their ability to displace the potent cannabinoid [3H] CP-55,940 (2) from its binding site in a membrane preparation from rat brain as described by Compton et al.42 Affinities for the CB2 receptor were determined by measuring the ability of the compounds to displace [3H] CP-55,940 from a cloned human receptor preparation using the procedure described by Showalter et al.17 The results of these determinations are summarized in Tables 2 and 3. Also included in Tables 2 and 3 are the receptor affinities for Δ9-THC (1) and WIN55,212-2 (4).
In the 4-alkyl-1-naphthoyl series (Table 2), the 4-methyl-1-propyl compounds, JWH-120 (41) and JWH-148, have very slight (Ki=1054±31 nM) and modest (Ki=123±8 nM) affinities respectively for the CB1 receptor. This pair of compounds is unusual in that the analog with an indole 2-methyl group (JWH-148) has significantly greater affinity for the CB1 receptor than the unsubstituted compound (JWH-120). The CB2 receptor affinities of these compounds are similar, for JWH-120 Ki=6.1±7 nM, and for JWH-148 Ki=14±1.0 nM. The 4-methyl-1-pentyl analogs, JWH-122 and JWH-149 have high affinities for both the CB1 and CB2 receptors. In particular, JWH-122, the compound lacking the indole C-2 methyl group, has subnanomolar affinity for the CB1 receptor (Ki=0.69±0.5 nM).24
The 4-ethyl- and 4-propyl-1-naphthoylindoles (Table 2) all have uniformly high affinities for both receptors. Following the usual trend for CB1 receptor affinities the 1-propyl compounds have lower affinities than the 1-pentyl analogs, and the compounds with an indole 2-methyl group have somewhat lower affinities than the unsubstituted analogs. The CB1 receptor affinities for these compounds range from Ki=0.46±0.03 nM for 1-pentyl-3-(4-ethyl-1-naphthoyl)indole, JWH-210, to Ki=70±0.8 nM for 2-methyl-1-propyl-3-(4-ethyl-1-naphthoyl)indole, JWH-211. The CB1 receptor affinities for the 3-(4-propyl-1-naphthoyl) indoles are comparable to those of the 4-ethyl analogs. The CB2 receptor affinities for these eight compounds fall within a narrow range, from Ki=0.62±0.04 for 2-methyl-1-pentyl-3-(4-propyl-1-naphthoyl)indole, JWH-181, to Ki=12±0.8 for 2-methyl-1-propyl-3-(4-ethyl-1-naphthoyl)indole, JWH-211.
The 4-butyl-1-naphthoylindoles (Table 1) have significantly lower affinities for the CB1 receptor than the 4-ethyl and 4-propyl-1-naphthoylindoles. In particular, the affinities of 1-propyl-3-(4-butyl-1-naphthoyl)indole, JWH-239 (Ki=342±20 nM) and its 2-methylindole analog, JWH-241 (Ki=147±20 nM) are significantly lower than the ethyl and propyl analogs. The CB1 receptor affinities of the 1-pentyl-3-(4-butyl-1-naphthoyl)indoles are much higher than those of the 1-propyl compounds. 1-Pentyl-3-(4-butyl-1-naphthoyl)indole, JWH-240 has Ki=14±1 nM and the 2-methylindole analog, JWH-242, has Ki=42±9 nM. Again, the values for the CB2 receptor affinities of the 4-butyl-1-naphthoylindoles are not nearly as diverse as the CB1 affinities. The 1-propyl analogs, JWH-239 and JWH-241 have Ki=52±6 nM and Ki=49±7 nM, respectively. The CB2 receptor affinities for the 1-pentyl compounds, JWH-240 and JWH-242 are essentially identical with Ki=7.2±13 nM for JWH-240 and Ki=6.5±0.3 nM for JWH-242.
We had previously reported the CB1 and CB2 receptor affinities of 2-methyl-1-propyl-3-(7-methyl-1-naphthoyl)indole (8), JWH-046 with Ki=343±38 nM at CB1 and Ki=16±5 nM at CB2 (Table 2), which is comparable in its affinity for both receptors and in CB2 selectivity to JWH-015 (6).21 JWH-076, the analog of JWH-046 lacking the 2-methyl group, has slightly greater affinity for the CB1 receptor than JWH-046 (Ki=214±11 nM), but considerably less affinity for the CB2 receptor (Ki=106±46 nM). 2-Methyl-1-pentyl-3-(7-methyl-1-naphthoyl) indole (9, JWH-048) was reported previously and has Ki=10.7±1 nM for the CB1 receptor and Ki=0.49±0.1 nM for the CB2 receptor.21 These values are very similar to those of the analog without the 7-methyl group, JWH-007 (5), which has Ki=9.5±4.5 nM at the CB1 receptor and Ki=2.9±2.6 nM at the CB2 receptor.19-21
For the 1-propyl-3-(7-ethyl-1-naphthoyl)indoles (Table 2), the 2-methyl compound, JWH-236 has considerably lower affinity than its 7-methyl homologue for both the CB1 and CB2 receptors with Ki=1351±204 nM at CB1 and Ki=240±63 nM at CB2. The analog lacking the 2-methyl group, JWH-235 has receptor affinities very similar to those of the 7 -methyl compound, with Ki=338±34 nM at CB1 and Ki=123±34 nM at CB2. The 1-pentyl compounds both have considerably greater affinities for both receptors than the 1-propyl analogs. For 1-pentyl-3-(7-ethyl-1-naphthoyl)indole, JWH-234, Ki=8.4±1.8 nM at CB1 and Ki=3.8±0.6 nM at CB2. The 2-methyl analog has slightly less affinity for both receptors with Ki=28±3 nM at CB1 and Ki=5.6±0.7 nM at CB2.
In the 3-(4-methoxy-1-naphthoyl)indole series the receptor affinities of all four of the 1-propyl- and 1-pentyl-3-(4-methoxy-1-naphthoyl)indoles have been reported previously (Table 2).21,24 The 1-propyl-2-methyl analog (JWH-094) has little affinity for the CB1 receptor with Ki=476±67 nM, and only modest affinity for the CB2 receptor (Ki=97±3 nM). However, JWH-079, 1-propyl-3-(4-methoxy-1-naphthoyl)indole has Ki=63±3 nM at CB1 and good affinity for the CB2 receptor (Ki=32±6 nM). The 1-pentyl analogs have considerably higher affinity for both receptors. 1-Pentyl-3-(4-methoxy-1-naphthoyl)indole (JWH-081) has very high affinity for the CB1 receptor (Ki=1.2±0.03 nM) and high affinity for the CB2 receptor (Ki=12.4±2.2 nM). The 2-methyl analog (JWH-098, 7) has slightly lower affinity for the CB1 receptor with Ki=4.5±0.1 nM and somewhat higher affinity for the CB2 receptor (Ki=1.9±0.3 nM).
The 3-(4-ethoxy-1-naphthoyl)indoles have from somewhat to considerably lower affinity for both receptors than the 4-methoxy analogs (Table 3). At the CB1 receptor the 1-propyl compounds have Ki=220±29 nM and Ki=767±105 nM, respectively, for 1-propyl-3-(4-ethoxy-1-naphthoyl)indole (JWH-259) and 2-methyl-1-propyl-3-(4-ethoxy-1-naphthoyl)indole (JWH-261). JWH-259 has modest affinity (Ki=74±7 nM), while the 2-methyl analog (JWH-261) has even less affinity (Ki=221±14 nM) at the CB2 receptor. The 1-pentyl compounds have considerably greater affinity for both receptors than the corresponding 1-propylindoles. 1-Pentyl-3-(4-ethoxy-1-naphthoyl)indole (JWH-258) has Ki=4.6±0.6 nM at CB1 and Ki=10.5 ±1.3 nM at the CB2 receptor. The 2-methyl compound, 2-methyl-1-pentyl-3-(4-ethoxy-1-naphthoyl)indole (JWH-260) has Ki=29±0.4 nM at CB1 and Ki=25±1.9 nM at the CB2 receptor.
None of the 1-alkyl-3-(2-methoxy-1-naphthoyl)indoles (JWH-265-JWH-268, Table 2) have significant affinity for the CB1 receptor, with Ki=381±16 nM for 1-pentyl-3-(2 -methoxy-1-naphthoyl)indole (JWH-267) to Ki=>10,000 nM for the 1-propyl-2-methyl analog (JWH-266). The CB2 receptor affinities of this series of compounds are considerably greater than for the CB1 receptor. 1-Propyl-3-(2-methoxy-1-naphthoyl)indole (JWH-265) has moderate affinity, Ki=80±13 nM, for the CB2 receptor while the 2-methyl analog (JWH-266) has little affinity for the receptor, Ki=455±55 nM. 1-Pentyl-3-(2-methoxy-1-naphthoyl)indole (JWH-267) has very high affinity, Ki=7.35 0.14 nM, for the CB2 receptor and the 2-methyl compound (JWH-268) has somewhat less affinity with Ki=40±1 nM.
The only 6-methoxy-1-naphthoylindole with significant affinity for the CB1 receptor is 1-pentyl-3-(6-methoxy-1-naphthoyl)indole (JWH-166, Table 2) which has good affinity with Ki=44±10 nM. The affinities of the other three members of this series for the CB1 receptor range from Ki=240±24 nM for 2-methyl-1-pentyl-3-(6-methoxy-1-naphthoyl)indole (JWH-153) to Ki=>10,000 for the 1-propyl-2-methyl analog (JWH-151, 42). The 1-pentyl-3-(6-methoxy-1-naphthoyl)indoles (JWH-153, JWH-166) have similar and high CB2 affinities with Ki=11±1 nM and Ki=1.9±0.1 nM, respectively. 1-Propyl-3-(6-methoxy-1-naphthoyl)indole (JWH-163) has Ki=138±12 nM at CB2, while the 2-methyl compound (JWH-151, 42) has significant affinity for the CB2 receptor, Ki=30±1 nM. JWH-151 is a highly selective (>333-fold) ligand for the CB2 receptor with effectively no affinity for the CB1 receptor.
In the 7-methoxy-1-naphthoyl series (Table 3), neither of the 1-propyl compounds has high affinity for either the CB1 or CB2 receptor (Table 3). 1-Propyl-3-(7-methoxy-1-naphthoyl)indole (JWH-165) has Ki=204±26 nM at the CB1 receptor and Ki=71±8 nM at the CB2 receptor. The 2-methyl analog (JWH-160) has Ki=1568±201 nM at CB1 and Ki=441 ±110 mM at CB2. In contrast, 1-pentyl-3-(7-methoxy-1-naphthoyl)indole (JWH-164) has Ki=6.6±0.7 nM at the CB1 receptor and Ki=6.9±0.2 nM at the CB2 receptor. The compound with the 2-methyl group, 2-methyl-1-pentyl-3-(7-methoxy-1-naphthoyl)indole (JWH-159), has somewhat less affinity for the CB1 receptor than JWH-164 with Ki=45±1 nM and similar affinity for the CB2 receptor, Ki=10.4±1.4 nM.
Three of the indole derivatives synthesized in the course of this SAR study are highly selective for the CB2 receptor. These are 1-propyl-3-(4-methyl-1-naphthoyl)indole, JWH-120 (41) which is 173-fold selective, 1-pentyl-3-(2-methoxy-1-naphthoyl)indole, JWH-267 (32), 53 -fold selective and 1-propyl-2-methyl-3-(6-methoxy-1-naphthoyl)indole, JWH-151 (42) which is >333 fold selective. In order to evaluate the efficacy of these compounds, their ability to stimulate GTPγS binding was determined. This is a functional assay which measures G-protein coupled receptor activation using [35S]GTPγS binding.43 Chinese Hamster Ovary (CHO) cells stably expressing the human CB2 receptor were employed in this determination (see Experimental). The results of these determinations are summarized in Table 4. The stimulation is normalized to that produced by 3 mM CP-55,940 (2), a maximally effective concentration of this standard cannabinoid agonist. In addition to indoles 32, 41 and 42 the [35S]GTPγS binding for JWH-015, 1-propyl-2-methyl-3-(1-naphthoyl)indole (6), the lead compound for the search for CB2 selective cannabimimetic indoles, was determined, and the data are included in Table 3. Also included in Table 4 are data for two dibenzopyran-based cannabinoids which are highly selective for the CB2 receptor, 1-deoxy-3-(1′,1′-dimethylbutyl)-Δ8-THC (JWH-133, 43) which has excellent affinity for the CB2 receptor (Ki=3.4±1.0 nM) and little affinity for the CB1 receptor (Ki=677±132 nM)44 and 1-methoxy-3-(1′,1′-dimethylhexyl)-Δ8-THC (JWH-229, 44) with Ki=18±2 nM at CB2 and Ki=3134±110 nM at CB1.45
As indicated in Table 4 all six of these compounds are potent in the [35S]GTPγS assay with EC50 values from 4.0±1.0 nM for JWH-133 (43) to 17.7±1.0 nM for JWH-015 (6). Two of these CB2 receptor ligands, JWH-133 (43) and 1-propyl-2-methyl-3-(6-methoxy-1-naphthoyl)indole, JWH-151 (42) are highly efficacious with Emax values of 111.5±13.6% and 108.5±13.0% respectively, relative to CP-55,940., The other three cannabimimetic indoles, 1-propyl-2-methyl-3-(1-naphthoyl)indole, JWH-015 (6), 1-propyl-3-(4-methyl-1-naphthoyl)indole, JWH-120 (41) and 1-pentyl-3-(2-methoxy-1-naphthoyl)indole, JWH-267 (32), plus 1-methoxy-3-(1′,1′-dimethylhexyl)-Δ8-THC, JWH-229 (44) are all partial agonists relative to CP-55,940 with Emax values from 65.7±6.4% (JWH-015) to 78.1±10.7% (JWH-120).
Conclusions
The CB1 receptor affinities for 1-pentyl- and 1-propyl-3-(1-naphthoyl)indoles with 4- and 7-alkylnaphthoyl and 2-, 4-methoxy, 6-, 7-methoxy and 4-ethoxynaphthoyl substituents indicate that receptor affinity is enhanced considerably by the presence of small alkyl groups (methyl, ethyl, propyl) at C-4. Methyl or ethyl substituents at C-7 of the naphthoyl group have little effect on affinity relative to the unsubstitued compounds. A methoxy substituent at C-4 enhances CB1 receptor affinity, while a 4-ethoxy substituent has relatively little effect on affinity. 6-Methoxy-1-naphthoylindoles have attenuated CB1 receptor affinities relative to the unsubstitued compounds, while a 7-methoxy substituent has little effect upon affinity. A 2 -methoxy substituent effectively destroys affinity for the CB1 receptor. These conclusions have been rationalized in terms of molecular modeling and receptor docking studies.
The CB2 receptor affinities of these naphthoylindoles show considerably less variation than their CB1 affinities. Only four compounds with alkylnaphthoyl substituents have CB2 receptor affinities greater than 52 nM. Three of these are 1-propyl-3-(7-methyl or 7-ethyl-1-naphthoyl)indoles and the fourth is 1-propyl-3-(-1-naphthoyl)indole. In the alkoxynaphthoyl series there is no clear pattern. Most of the alkoxynaphthoyl cannabimimetic indoles have good to moderate affinity for the CB2 receptor, however a 1-propyl-2-methylindole substitution pattern leads to attenuated affinity in several cases.
A 2-methyl substituent on the indole nucleus results in a decrease in affinity for either receptor relative to the unsubstituted analog. Similarly, the 1-propylindoles in general have lower affinities at both receptors than the 1-pentyl compounds. However, the differences in CB2 receptor affinities as a function of N-substitution are less pronounced than is the case with CB1 affinities. Although three new cannabimimetic indoles with potentially useful CB2 selectivity have been identified, there do not appear to be sufficient data to permit the rational design of additional indole based CB2 selective ligands.
Experimental
General. IR spectra were obtained using Nicolet 5DX or Magna spectrometers; 1H and 13C NMR spectra were recorded on a Bruker 300AC spectrometer. Mass spectral analyses were performed on a Hewlett-Packard 5890A capillary gas chromatograph equipped with a mass sensitive detector. HRMS data were obtained in the Mass Spectrometry Laboratory, School of Chemical Sciences, University of Illinois. Ether and THF were distilled from Nabenzophenone ketyl immediately before use, and other solvents were purified using standard procedures. Column chromatography was carried out on Sorbent Technologies silica gel (32-63 μ) using the indicated solvents as eluents. All new compounds were homogeneous to TLC and 13C NMR. All target compounds were homogeneous to GLC or TLC in two different solvent systems. TLC was carried out using 200 mm silica gel plates using the indicated solvents. GLC analyses were performed on the Hewlett-Packard 5890A GC/MS using a 60 m carbowax column and helium gas as a carrier. An initial column temperature of 60° C. was employed and the temperature was increased at a rate of 1.5° C./min to a maximum temperature of 300° C. with a total run time of 20 min. Elemental analyses were performed by Atlantic Microlab, Norcross, Ga.
1-Propyl-3-(4-methyl-1-naphthoyl)indole JWH-120 (41, Method B). To a stirred solution of 0.080 g (0.50 mmol) of 1-propylindole in 1.5 mL of dry CH2Cl2 at 0° C. under N2 was added dropwise 0.75 mL (0.75 mmol) of Me2AlCl (1 M in hexanes). The solution was stirred at 0° C. for 30 min and a solution of 0.122 g (0.6 mmol) of freshly prepared 4-methyl-1-naphthoyl chloride in 1.5 mL of CH2Cl2. The acid chloride was prepared form 0.122 g (0.60 mmol) of 4-methyl-1-naphthoic acid to which was added dropwise 1 mL of thionyl chloride at a rate sufficient to maintain a steady evolution of gas. This solution was heated at reflux for 15 min, cooled to ambient temperature and the excess thionyl chloride was removed in vacuo to give the acid chloride which was used without further purification. The deep red acylation reaction mixture was stirred at 0° C. until the reaction was complete as indicated by tlc (approximately 1 h). The reaction mixture was poured carefully into iced 1 M aqueous HCl and extracted with three portions of CH2Cl2. The combined extracts were washed wth three portions of aqueous NaHCO3, dried (MgSO4) and the solvent was removed in vacuo to give the crude product. After chromatography (petroleum ether:ethyl acetate, 9: 1) there was obtained 0.140 g (86%) of indole 41 as a white solid. Recrystallization from hexanes/ethyl acetate gave the analytical sample: m.p. 207-208° C.; 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=7.3 Hz, 3H), 1.81-1.88 (m, 2H), 2.78 (s, 3H), 4.03 (t, J=7.0 Hz, 2H), 7.37-7.38 (m, 5H), 7.48-7.59 (m, 3H), 8.09 (d, J=8.2 Hz, 1H), 8.27 (d, J=8.2 Hz, 1H), 8.52-8.54 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.3, 19.7, 23.1, 48.7, 109.9, 117.6, 122.7, 122.9, 123.5, 124.2, 125.5, 125.8, 126.1, 126.3, 126.6, 127.0, 130.9, 132.8, 136.6, 137.0, 137.5, 137.9, 192.2; Anal. Calcd for C23H21NO: C, 84.37; H, 6.46; N, 4.28; Found: C, 84.25; H, 6.46; N, 4.30.
2-Methyl-1-propyl-3-(4-methyl-1-naphthoyl)indole JWH-148 (Method A). To a stirred solution of 14.1 mL (42.3 mmol) of 3.0 M MeMgBr in ether, diluted with 20 mL of THF, at 0° C. was added 4.62 g (35.2 mmol) of 2-methylindole in 10 mL of THF. The reaction mixture was allowed to warm to ambient temperature and a solution of 4-methyl-1-naphthoyl chloride (from 6.0 g, 35 mmol, of 4-methyl-1-naphthoic acid) in 5 mL of THF was added dropwise. The mixture was heated at reflux for 1.5 h and the reaction was quenched by the cautious addition of saturated aqueous NH4Cl. Stirring was continued until the solid precipitate was broken up and the solid was collected, and suspended in 200 mL of methanol. A solution of 3 g of KOH in 20 mL of water was added and the mixture was heated at reflux for 4 h. The precipitate was filtered off and dried to give 3.7 g (35%) of 2-methyl-3-(4-methyl-1-naphthoyl)indole was added to a stirred suspension of 1.48 g of powdered KOH in 5 mL of DMSO and 1.44 g (11.7 mmol) of 1-bromopropane was added. The reaction mixture was stirred at 80° C. for 18 h, poured into water and extracted with three portions of ethyl acetate. The combined extracts were washed with water, dried (Na2SO4) and the solvent was removed to give the crude product. Chromatography (petroleum ether:ethyl acetate, 10:1) gave 1.62 g (81%) of JWH-148 as a white solid: m.p. 135-137°; 1H NMR (300 MHz, CDCl3) δ 0.95 (t, J=7.4 Hz, 3H), 1.79 (sextet, J=7.4 Hz, 2H), 2.44 (s, 3H), 2.74 (s, 3H), 4.04 (t, J=7.3 Hz, 2H), 6.98 (t, J=7.9 Hz, 1H), 7.15 (t, J=7.9 Hz, 1H), 7.21-7.52 (m, 6H), 8.06 (d, J=8.5 Hz, 1H), 8.18 (d, J=8.3 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.6, 12.7, 20.0, 23.0, 44.9, 109.6, 115.2, 121.4, 121.9, 122.2, 124.4, 125.9, 126.2, 126.4, 126.6, 127.3, 130.6, 133.0, 136.2, 136.8, 139.0, 145.5, 193.7; MS (EI) m/z 341 (80), 326 (43), 324 (25), 141 (100); Anal. Calcd for C24H23NO: C, 84.42; H, 6.79; N, 4.10; Found: C, 84.39;H, 6.83; N, 4.08.
4-Ethyl-1-N,N-dipehylamidonaphthalene (19, R═C2H5). A mixture of 3.00 g, (19.2 mmol) of 1-ethylnaphthalene, 4.45 g (19.2 mmol) of diphenylcarbamyl chloride and 2.82 g (21.1 mmol) of anhydrous AlCl3 in 38.4 mL of 1,2-dichloroethane was heated at reflux for 6 h. After cooling, the reaction was poured onto a mixture of ice and concentrated HCl and extracted with ether. The extracts were dried (MgSO4) and the solvent was removed in vacuo. The residue was chromatographed (petroleum ether:ethyl acetate, 9:1) to give a white solid. Recrystallization gave 1.72 g (80%) of amide which used in the next step without further purification: m.p. 126-127°; 1H NMR (300 MHz, CDCl3) δ 1.29 (t, J=7.5 Hz, 3H), 3.02 (q, J=7.5 Hz, 2H), 7.08-7.31 (m, 12H), 7.49-7.57 (m, 2H), 8.02 (d, J=7.6 Hz, 1H), 8.32 (d, J=7.6 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 14.7, 26.0, 123.4, 124.0, 125.9, 126.1, 126.4, 127.2, 128.9, 131.0, 131.8, 132.6, 142.2, 143.2, 170.6; MS (EI) m/z 351 (12), 183 (100).
4-Ethyl-1-naphthoic acid (15). A mixture of 3.70 g (10.5 mmol) of the above diphenylamide, 23.2 g of KOH and 154 mL of diethylene glycol was heated at reflux for 8 h. The reaction mixture was cooled, diluted with 1.13 L of water and the precipitated solids filtered off. The filtrate was acidified with conc. HCl until precipitation was complete. The solid was collected and dried to give 1.90 g (90%) of acid 15: m.p. 125-127° C. (lit mp 129-130° C., ref. 26): 1H NMR (300 MHz, CDCl3) δ 1.44 (t, J=7.3 Hz, 3H), 3.20 (q, J=7.5 Hz, 2H), 7.44 (d, J=7.6 Hz, 1H), 7.59-7.70 (m, 2H), 8.16 (d, J=8.2 Hz, 1H), 8.39 (d, J=7.4 Hz, 1H), 9.21 (d, J=8.6 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) 614.7, 26.5, 123.8, 124.1, 125.0, 126.1, 126.6, 127.5, 131.9, 132.1, 147.5, 173.7
1-Propyl-3-(4-ethyl-1-naphthoyl)indole, JWH-212 (Method C). 4-Ethyl-1-naphthoic acid was converted to the acid chloride by the procedure described above in the preparation of 2-methyl-1-propyl-3-(4-methyl-1-naphthoyl)indole (JWH-148). To a solution of 4-ethyl-1-naphthoyl chloride, from 0.20 g (1.00 mmol) of 4-ethyl-1-naphthoic acid in 4.0 mL of toluene was added a solution of 0.19 g (1.20 mmol) of 1-propylindole. The solution was cooled to 0° C. and 0.26 mL (1.5 mmol) of 1.0 M EtAlCl2 in hexanes was added dropwise with stirring. The reaction mixture was warmed to ambient temperature and stirred for four days. After the addition of 2.0 mL of water, the layers were separated and the aqueous layer was extracted with ethyl acetate. The combined organic extracts were washed with water, dried (MgSO4) and concentrated in vacuo. The residue was chromatographed (petroleum ether:ether, 9:1) to give 0.190 g (56%) of JWH-212 as a yellow gum: 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=7.4 Hz, 3H), 1.45 (t, J=7.5 Hz, 3H), 1.78-1.90 (m, 2H), 3.19 (q, J=7.4 Hz, 2H), 4.04 (t, J=7.1 Hz, 2H), 7.35-7.40 (m, 5H), 7.44-7.53 (m, 2H), 7.55-7.61 (m, 2H), 8.14 (d, J=8.4 Hz, 1H), 8.26 (d, J=8.3 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) d 11.3, 14.9, 23.1, 26.1, 48.7, 109.9, 117.7, 122.7, 122.9, 123.5, 123.8, 124.6, 125.9, 126.0, 126.2, 126.8, 127.1, 131.1, 132.0, 137.0, 137.4, 137.9,142.5, 192.6; MS (EI) m/z 341 (100), 324 (65), 312 (91); HRMS Calcd for C24H23NO: 341.1780, Found 341.1779.
1-Pentyl-3-(4-ethyl-1-naphthoyl)indole, JWH-210. JWH-210 was prepared from 4-ethyl-1-naphthoic acid and 1-pentylindole by method C. From 0.20 g (1.00 mmol) of 4-ethyl-1-naphthoic acid and 0.22 g (1.20 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.193 g (52%) of JWH-210 as a viscous oil: 1H NMR (300 MHz, CDCl3) δ 0.88 (t, J=6.9 Hz, 3H), 1.24-1.35 (m, 3H), 1.40-1.49 (m, 4H), 1.76-1.85 (m, 2H), 3.21 (q, J=7.5 Hz, 2H), 4.05 (t, J=7.2 Hz, 2H), 7.35-7.42 (m, 5H), 7.49-7.52 (m 2), 7.55-7.64 (m, 2H), 8.16 (d, J=8.3 Hz, 1H), 8.32 (d, J=8.5 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) 13.8, 14.8,22.1,26.1,28.8,29.4,45.0, 109.9, 117.5, 122.6, 122.8, 123.4, 123.7, 124.0, 125.8, 126.0, 126.1, 126.7, 127.0, 131.1, 131.9, 136.9, 137.4, 137.8, 142.4, 192.2; MS (EI) m/z 369 (100), 352 (64), 312 (88); HRMS Calcd for C26H27NO: 369.2093, Found 369.2090.
2-Methyl-1-propyl-3-(4-ethyl-1-naphthoyl)indole, JWH-211. JWH-211 was prepared from 4-ethyl-1-naphthoic acid and 2-methyl-1-propylindole by method C. From 0.20 g (1.00 mmol) of 4-ethyl-1-naphthoic acid and 0.21 g (1.20 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.192 g (52%) of JWH-211 as a gum: 1H NMR (300 MHz, CDCl3) δ 1.00 (t, J=7.4 Hz, 3H), 1.43 (t, J=7.5 Hz, 3H), 1.78-1.90 (m, 2H), 2.48 (s, 3H), 3.20 (q, J=7.5 Hz, 2H), 4.10 (t, J=7.5 Hz, 2H), 7.00 (d, J=7.2 Hz, 1H), 7.15-7.25 (m, 2H), 7.30-7.35 (m, 2H), 7.40-7.55 (m, 3H), 8.14 (d, J=8.4 Hz, 1H), 8.18 (d, J=8.6 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.4, 12.5, 14.9, 22.9, 26.2, 44.8, 109.4, 115.1, 121.3, 121.8, 122.1, 123.8, 124.0, 125.8, 126.0, 126.3, 126.4, 127.1, 130.7, 132.1, 136.1, 138.8, 142.7, 145.4, 192.9; MS (EI) m/z 355 (100), 338 (40), 326 (96), 298 (38); HRMS Calcd for C25H25NO: 354.1858, Found 354.1852.
2-Methyl-1-pentyl-3-(4-ethyl-1-naphthoyl)indole, JWH-213. JWH-213 was prepared from 4-ethyl-1-naphthoic acid and 2-methyl-1-propylindole by method C. From 0.20 g (1.00 mmol) of 4-ethyl-1-naphthoic acid and 0.24 g (1.20 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.205 g (55%) of JWH-213 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.98 (t, J=7.0 Hz, 3H), 1.22-1.34 (t, J=7.3 Hz, 3H), 1.42-1.49 (m, 4H), 1.84-1.94 (m, 2H), 2.56 (s, 3H), 3.28 (q, J=7.5 Hz, 2H), 4.20 (t, J=7.5 Hz, 2H), 7.10 (d, J=7.3 Hz, 1H), 7.27-7.34 (m, 2H), 7.38-7.54 (m, 2H), 7.59-7.64 (m, 3H), 8.22 (d, J=8.4 Hz, 1H), 8.26 (d, J=8.2 Hz,1H); 13C NMR (75.5 MHz, CDCl3) δ 12.4, 13.8, 14.9, 22.3, 26.1,29.0,29.2,43.2, 109.3, 115.0, 121.2, 121.7, 122.0, 123.8, 123.9, 125.7, 125.9, 126.2, 126.3, 127.1, 130.7, 132.0, 136.0, 138.8, 142.5, 145.2, 193.6; MS (EI) m/z 383 (100), 368 (59), 354 (71), 298 (43); HRMS Calcd for C27H29NO: 383.2249, Found 383.2246.
4-Propyl-1-naphthoic acid (16). Acid 16 was prepared in a manner analogous to that employed for 4-ethyl-1-naphthoic acid: m.p. 134-135° C. (lit mp 141-142° C., ref. 26); 1H NMR (300 MHz, CDCl3) 67 1.06 (t, J=7.3 Hz, 3H), 1.78-1.86 (m, 2H), 3.13 (t, J=7.7 Hz, 2H), 7.41 (d, J=7.5 Hz, 1H), 7.56-7.67 (m, 2H), 8.14 (d, J=8.2 Hz, 1H), 8.34 (d, J=7.5 Hz, 1H), 9.16 (d, J=8.6 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 24.2, 23.8,35.7, 123.9, 124.3, 124.9, 126.0, 126.6, 127.5, 131.6, 132.1, 132.3, 146.1, 172.9; MS (ED) m/z 214 (59), 185 (100), 157 (20).
1-Propyl-3-(4-propyl-1-naphthoyl)indole, JWH-180. JWH-180 was prepared from 4-propyl-1-naphthoic acid and 1-propylindole by method C. From 0.11 g (0.51 mmol) of 4-propyl-1-naphthoic acid and 0.098 g (0.62 mmol) of 1-propylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.098 g (54%) of JWH-180 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=7.4 Hz, 3H), 1.08 (t, J=7.3 Hz, 3H), 1.80-1.88 (m, 4H), 3.12 (t, J=7.7 Hz, 2H), 4.05 (t, J=7.1 Hz, 2H), 7.34-7.43 (m, 5H), 7.46-7.50 (m,2H), 7.52-7.60 (m, 2H), 8.12 (d, J=8.1 Hz, 1H), 8.24 (d, J=8.1 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.2, 14.3, 23.1, 23.8, 35.4, 48.7, 109.9, 117.6, 122.7, 122.9, 123.5, 124.0, 124.6, 125.7, 126.0, 126.1, 126.7, 127.0, 131.2, 132.2, 137.0, 137.5, 137.9, 141.0, 192.0; MS (EI) m/z 355 (100), 338 (57), 326 (52); HRMS Calcd for C25H25NO: 355.1936, Found 355.1924.
1-Pentyl-3-(4-propyl-1-naphthoyl)indole, JWH-182. JWH-182 was prepared from 4-propyl-1-naphthoic acid and 1-pentylindole by method C. From 0.11 g (0.51 mmol) of 4-propyl-1-naphthoic acid and 0.11 g (0.62 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.099 g (50%) of JWH-182 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.86 (t, J=6.8 Hz, 3H), 1.09 (t, J=7.3 Hz, 3H), 1.24-1.32 (m, 4H), 1.79-1.92 (m, 4H), 3.13 (t, J=7.7 Hz, 2H), 4.07 (t, J=7.2 Hz, 2H), 7.35-7.44 (m, 5H), 7.46-7.50 (m, 2H), 7.52-7.60 (m, 2H), 8.13 (d, J=8.4 Hz, 1H), 8.26 (d, J=8.2 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 13.8, 14.3, 22.1, 23.8, 28.9, 29.5, 35.4, 47.1, 109.9, 117.6, 122.7, 122.9, 123.5, 124.0, 124.6, 125.7, 126.0, 126.1, 126.7, 127.0, 131.2, 132.2, 137.0, 137.5, 137.8, 141.0, 192.0; MS (EI) m/z 383 (100), 366 (63), 326 (75); HRMS Calcd for C27H29NO: 383.2249, Found 383.2243.
2-Methyl-1-propyl-3-(4-propyl-1-naphthoyl)indole, JWH-189. JWH-189 was prepared from 4-propyl-1-naphthoic acid and 2-methyl-1-propylindole by method C. From 0.12 g (0.56 mmol) of 4-propyl-1-naphthoic acid and 0.12 g (0.67 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.12 g (60%) of JWH-189 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.97-1.08 (m, 6H), 1.80-1.88 (m, 4H), 2.48 (s, 3H), 3.13 (t, J=7.6 Hz, 2H), 4.10 (t, J=7.5 Hz, 2H), 7.00 (d, J=7.6 Hz, 1H), 7.15-7.25 (m, 2H), 7.30-7.38 (m, 2H), 7.40-7.56 (m, 3H), 8.13 (d, J=8.5 Hz, 1H), 8.25 (d, J=8.3 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.4, 12.9, 14.2, 22.9, 23.8, 35.4, 44.8, 109.4, 121.3, 122.0, 122.1, 122.9, 124.1, 125.2, 125.7, 126.0, 126.4, 127.4, 128.0, 130.9, 132.3, 136.0, 139.1, 141.1, 145.7, 192.9; MS (EI) m/z 369 (100), 352 (40), 326 (87); HRMS Calcd for C26H27NO: 369.2093, Found 369.2088.
2-Methyl-1-pentyl-3-(4-propyl-1-naphthoyl)indole, JWH-181. JWH-181 was prepared from 4-propyl-1-naphthoic acid and 2-methyl-1-pentylindole by method C. From 0.11 g (0.51 mmol) of 4-propyl-1-naphthoic acid and 0.12 g (0.62 mmol) of 2-methyl-1-pentylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.11 g (54%) of JWH181 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.93 (t, J=7.4 Hz, 3H), 1.05 (t, J=7.3 Hz, 3H), 1.36-1.40 (m, 4H), 1.80-1.87 (m, 4H), 2.48 (s, 3H), 3.13 (t, J=7.6 Hz, 2H), 4.12 (t, J=7.6 Hz, 2H), 7.00 (d, J=7.5 Hz, 1H), 7.15-7.22 (m, 2H), 7.30-7.35 (m, 2H), 7.40-7.55 (m, 3H), 8.13 (d, J=8.5 Hz, 1H), 8.17 (d, J=8.4 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 12.5, 13.9, 14.2, 22.4, 23.8, 29.1, 29.3, 35.4, 43.3, 109.3, 115.1, 121.3, 121.8, 122.1, 124.1, 125.2, 125.6, 126.3, 126.4, 127.1, 130.8, 132.2, 136.0, 138.8, 141.0, 145.3, 193.8; MS (EI) m/z 397 (100), 382 (56), 354 (81); HRMS Calcd for C28H31NO: 397.2406, Found 397.2394.
4-Butyl-1-naphthoic acid (17). Acid 17 was prepared in a manner analogous to that employed for 4-ethyl-1-naphthoic acid: m.p. 136-137° C. (lit mp 148-148.5° C., ref. 26); 1H NMR (300 MHz, CDCl3) δ 0.99 (t, J=7.3 Hz, 3H), 1.42-1.54 (m, 2H), 1.73-1.81 (m, 2H), 3.14 (t, J=7.7 Hz, 2H), 7.41 (d, J=7.3 Hz, 1H), 7.56-7.67 (m, 2H), 8.14 (d, J=8.2 Hz, 1H), 8.33 (d, J=7.5 Hz, 1H), 9.15 (d,J=8.2 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 14.0, 22.9, 32.8, 33.4, 123.9, 124.3, 124.9, 126.0, 126.6, 127.5, 131.6, 132.1, 132.3, 146.3, 172.9; MS (EI) m/z 228 (53), 185 (100), 157 (36).
1-Propyl-3-(4-butyl-1-naphthoyl)indole, JWH-239. JWH-239 was prepared from 4-butyl-1-naphthoic acid and 1-propylindole by method C. From 0.18 g (0.79 mmol) of 4-butyl-1 -naphthoic acid and 0.15 g (0.95 mmol) of 1-propylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.19 g (62%) of JWH-239 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=7.4 Hz, 3H), 1.02 (t, J=7.3 Hz, 3H), 1.46-1.58 (m, 2H), 1.71-1.88 (m, 4H), 3.15 (t, J=7.8 Hz, 2H), 4.04 (t, J=7.1 Hz, 2H), 7.35-7.43 (m, 5H), 7.46-7.50 (m, 2H), 7.52-7.60 (m, 2H), 8.14 (d, J=8.3 Hz, 1H), 8.27 (d, J=8.2 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.3, 14.0, 22.9, 23.1, 32.9, 33.1, 48.7, 109.9, 117.6, 122.7, 122.9, 123.5, 124.0, 124.5, 125.7, 126.0, 126.2, 126.7, 127.0, 131.2, 132.1, 137.0, 137.4, 137.9, 141.3, 192.3; MS (EI) m/z 369 (100), 352 (70), 340 (53); HRMS Calcd for C26H27NO: 369.2093, Found 369.2094.
1-Pentyl-3-(4-butyl-1-naphthoyl)indole, JWH-240. JWH-240 was prepared from 4-butyl-1-naphthoic acid and 1-pentylindole by method C. From 0.17 g (0.75 mmol) of 4-butyl-1-naphthoic acid and 0.17 g (0.89 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.17 g (58%) of JWH-240 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.87 (t, J=6.8 Hz, 3H), 1.03 (t, J=7.3 Hz, 3H), 1.24-1.35 (m, 4H), 1.47-1.59 (m, 2H), 1.76-1.86 (m, 4H), 3.16 (t, J=7.8 Hz, 2H), 4.06 (t, J=7.2 Hz, 2H), 7.34-7.42 (m, 5H), 7.45-7.53 (m, 2H), 7.56-7.60 (m, 2H), 8.15 (d, J=8.4 Hz, 1H), 8.28 (d, J=8.3 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 13.8, 14.0, 22.1, 22.9, 28.8, 29.4, 32.9, 33.1, 47.0, 109.9, 117.6, 122.7, 122.9, 123.4, 124.0, 124.5, 125.7, 126.0, 126.1, 126.7, 127.0, 131.2, 132.1, 137.0, 137.4, 137.8, 141.3, 192.3; MS (EI) m/z 397 (100), 380 (59), 340 (82); HRMS Calcd for C29H31NO: 397.2406, Found 397.2400.
2-Methyl-1-propyl-3-(4-butyl-1-naphthoyl)indole, JWH-241. JWH-241 was prepared from 4-butyl-1-naphthoic acid and 2-methyl-1-propylindole by method C. From 0.18 g (0.79 mmol) of 4-butyl-1-naphthoic acid and 0.16 g (0.95 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.18 g (62%) of JWH-241 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.97-1.03 (m, 6H), 1.44-1.56 (m, 2H), 1.77-1.90 (m, 4H), 2.49 (s, 3H), 3.16 (t,J=7.7 Hz, 2H), 4.13 (t,J=7.9 Hz, 2H), 7.02 (d,J=7.4 Hz, 1H), 7.16-7.25 (m, 2H), 7.30-7.36 (m, 2H), 7.42-7.56 (m, 3H), 8.13 (d, J=8.5 Hz, 1H), 8.25 (d, J=8.3 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.4, 12.5, 14.0, 22.8, 22.9, 32.9, 33.1, 44.8, 109.4, 115.0, 121.3, 121.7, 122.1, 124.0, 125.0, 125.7, 125.9, 126.2, 126.4, 127.1, 130.8, 132.2, 136.1, 138.8, 141.3, 145.4, 193.7; MS (EI) m/z 383 (100), 366 (34), 326 (95); Calcd for C27H29NO: 383.2249, Found 383.2246.
2-Methyl-1-pentyl-3-(4-butyl-1-naphthoyl)indole, JWH-242. JWH-242 was prepared from 4-butyl-1-naphthoic acid and 2-methyl-1-pentylindole by method C. From 0.17 g (0.75 mmol) of 4-butyl-1-naphthoic acid and 0.18 g (0.89 mmol) of 2-methyl-1-pentylindole there was obtained after chromatography (petroleum ether:ether, 9:1) 0.17 g (56%) of JWH-242 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.92 (t, J=6.9 Hz, 3H), 1.00 (t, J=7.3 Hz, 3H), 1.39-1.45 (m, 4H), 1.47-1.55 (m, 2H), 1.74-1.84 (m, 4H), 2.48 (s, 3H), 3.16 (t, J=7.7 Hz, 2H), 4.12 (t, J=7.6 Hz, 2H), 7.02 (t, J=7.4 Hz, 1H), 7.16-7.26 (m, 2H), 7.32-7.36 (m, 2H), 7.40-7.56 (m, 3H), 8.14 (d, J=8.5 Hz, 1H), 8.18 (d, J=8.5 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 12.5, 13.9, 14.0, 22.4, 22.8, 29.1, 29.3, 32.9, 33.1, 43.3, 109.3, 121.3, 121.8, 122.1, 124.0, 125.1, 125.7, 125.9, 126.2, 126.4; MS (EI) m/z 411 (89), 396 (60), 354 (100); Calcd for C29H33NO: 411.2562, Found 411.2563.
1-Propyl-3-(7-methyl-1-naphthoyl)indole, JWH-076 (Method A). JWH-076 was prepared from 1-propylindole and 7-methyl-1-naphthoyl chloride by method B. From 0.186 g (0.65 mmol) of 3-(7-methyl-1-naphthoyl)indole and 0.5 mL (5.5 mmol) of 1-bromopropane there was obtained after chromatography (petroleum ether:ethyl acetate, 1:3) 0.207 g (97%) of JWH-076 as a tan gum: 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=7.3 Hz, 3H), 1.85 (q, J=7.3 Hz, 2H), 2.44 (s, 3H), 4.05 (t, J=7.2 Hz, 2H), 7.35-7.47 (m, 6H), 7.62 (d, J=6.8 Hz, 1H), 7.81 (d, J=8.3 Hz, 1H), 7.92 (d, J=8.1 Hz, 1H), 7.98 (s, 1H), 8.52 (t, J=5.4 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.3, 21.9, 23.1, 48.7, 110.0, 117.6, 122.8, 123.0, 123.5, 124.8, 125.9, 127.0, 127.9, 128.6, 129.7, 130.9, 132.0, 136.7, 137.0, 138.0, 138.4, 192.3; MS (EI) m/z 327 (82), 310 (43), 298 (58), 186 (100).
7-Ethyl-1-naphthoic Acid (24). To a mixture of 9.56 g of AlCl3(64 mmol) and 4.05 mL of acetyl chloride (57 mmol) was added, with stirring, 21.3 mL of 1,2-dichloroethane. To this light brown solution was added dropwise a solution of 10.0 g (64 mmol) of 2-ethylnaphthalene in 7.1 mL of 1,2-dichloroethane. The reaction mixture was allowed to stand at ambient temperature for 18 h, diluted with dichloromethane, and shaken with dilute aqueous HCl. After washing with saturated aqueous NaHCO3 and water, the organic phase was dried (MgSO4) and the solvents were removed in vacuo. Chromatography (petroleum ether:ethyl acetate, 95:5) gave 9.88 g (86%) of 1-acetyl-7-ethylnaphthalene (26) as a colorless oil: 1H NMR (300 MHz, CDCl3) δ 1.33 (t, J=7.6 Hz, 3H), 2.72 (s, 3H), 2.83 (q, J=7.5 Hz, 2H), 7.39-7.50 (m, 2H), 7.79 (d, J=8.4 Hz, 1H), 8.56 (s, 1H); 13C NMR (75.5 MHz, CDCl3) δ 15.5, 29.5, 30.0, 123.4, 123.8, 127.5, 128.3, 128.7, 130.3, 132.4, 132.7, 134.6, 144.3, 202.1.
A solution of 8.7 g (44 mmol) of iodine in 15 mL of pyridine was added to 8.70 g (44 mmol) of 1-acetyl-7-ethylnaphthalene, and the solution was stirred at 100° C. for 30 min. After cooling to ambient temperature the precipitated pyridinium salt was suspended in methanol and collected by filtration. After drying in vacuo at 45° C. there was obtained 23.5 g of salt 27 as a brown solid, which was added to 250 mL of water without further purification and 20 g of solid NaOH were added. The reaction mixture was heated at reflux for 2 h, cooled to ambient temperature and acidified with 10% aqueous HCl to give 7.56 g of acid 24 as dark purple crystals. Recrystallization from aqueous methanol gave 5.74 g (65%) of product as a brown solid. For further purification, to a solution of 1.0 g of acid in 15 mL of methanol was added 3 mL of H2SO4 and the reaction mixture was heated at reflux for 2 h. The reaction mixture was concentrated in vacuo, diluted with ether, washed with water and dried (MgSO4). The solvent was removed in vacuo to give a brown oil, which was chromatographed (petroleum ether:ethyl acetate, 95:5) to give 0.42 g (42%) of ester as a colorless oil: 1H NMR (300 MHz, CDCl3) δ 1.29 (t, J=7.6 Hz, 3H), 2.84 (q, J=7.3 Hz, 2H), 3.98 (s, 3H), 7.39 (t, J=7.6 Hz, 1H), 7.38 (d, J=8.0 Hz, 1H), 7.77 (d, J=8.4 Hz, 1H), 7.94 (d, J=8.1 Hz, 1H), 8.07 (d, J=7.7 Hz, 1H), 8.74 (s, 1H); 13C NMR (75.5 MHz, CDCl3) δ 15.5, 29.5, 51.9, 123.5, 126.3, 127.2, 128.4, 130.2, 131.6, 132.3, 133.0, 143.9, 168.0.
To a suspension of 1.66 g (8.4 mmol) of ester in 50 mL of water was added 10 g of KOH and the mixture was heated at reflux for 18 h. After acidification with conc. HCl to pH 5 the precipitated solid was filtered out to give, after drying in vacuo, 1.32 g (80%) of 7-ethyl-1-naphthoic acid (24) as an off white solid, which was used in subsequent steps without further purification. Recrystallizatiion from petroleum ether gave fine white needles: mp 117-120° C. (lit. mp 126° C., ref. 49): 1H NMR (300 MHz, CDCl3) δ 1.37 (t, J=7.5 Hz, 3H), 2.90 (q, J=7.5 Hz, 2H), 7.44-7.51 (m, 2H), 7.85 (d, J=8.4 Hz, 1H), 8.06 (d, J=8.2 Hz, 1H), 8.38 (dd, J=1.0, 7.2 Hz, 1H), 8.88 (s, 1H).
1-Propyl-3-(7-ethyl-1-naphthoyl)indole, JWH-235. JWH-235 was prepared from 7-ethyl-1-naphthoic acid by method B. From 0.150 g (0.75 mmol) of 7-ethyl-1-naphthoic acid and 0.08 g (0.50 mmol) of 1-propylindole there was obtained 0.150 g (88%) of JWH-235 as a pale brown oil after chromatography (petroleum ether:ethyl acetate, 95:5): 1H NMR (300 MHz, CDCl3) δ 0.83 (t, J=7.3 Hz, 3H), 1.21 (t, J=7.5 Hz, 3H), 1.70-1.83 (m, 2H), 2.72 (q, J=7.5 Hz, 2H), 3.95 (t, J=7.1 Hz, 2H), 7.20-7.38 (m, 5H), 7.42 (d, J=7.2 Hz, 1H), 7.58 (dd, J=7.2, 0.8 Hz, 1H), 7.80 (d, J=8.3 Hz, 1H), 7.88 (d, J=8.3 Hz, 1H), 8.01 (s, 1H), 8.53-8.56 (m 1); 13C NMR (75.5 MHz, CDCl3) δ 11.2, 15.6, 22.9, 29.2, 48.6, 110.0, 117.4, 122.7, 122.8, 123.4, 123.5, 123.6, 125.9, 126.9, 127.3, 128.1, 129.6, 130.9, 132.2, 137.0, 138.0, 138.4, 142.8, 192.1; MS (EI) m/z 341 (100), 312 (80); HRMS Calcd for C24H23NO: 341.1760, Found 341.1779.
1-Pentyl-3-(7-ethyl-1-naphthoyl)indole, JWH-234. JWH-234 was prepared from 7-ethyl-1-naphthoic acid and 1-pentylindole by method B. From 0.150 g (0.75 mmol) of 7-ethyl-1-naphthoic acid and 0.094 g (0.5 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 95:5) 0.100 g (54%) of JWH-234 as a pale brown oil: 1H NMR (300 MHz, CDCl3) δ 0.85 (t, J=7.0 Hz, 3H), 1.21-1.46 (m, 7H), 1.75-1.84 (m, 2H), 2.73 (q, J=7.6 Hz, 2H), 4.05 (t, J=7.2 Hz, 2H), 7.34-7.41 (m, 5H), 7.45 (d, J=7.2 Hz, 1H), 7.61 (dd, J=7.2, 1.0 Hz, 1H), 7.83 (d, J=8.3 Hz, 1H), 7.92 (d, J=8.3 Hz, 1H), 8.00 (s, 1H), 8.51-8.53 (m,1H); 13C NMR (75.5 MHz, CDCl3) δ 13.8, 15.7, 22.1, 28.7, 29.3, 29.5, 47.1, 109.9, 117.6, 122.8, 122.9, 123.5, 123.6, 123.7, 126.0, 127.0, 127.4, 128.1, 129.7, 131.0, 132.3, 137.0, 137.9, 138.5, 143.0, 192.2; MS (EI) m/z 369 (75), 352 (40), 312 (50), 207 (100); HRMS Calcd for C26H27NO: 369.2091, Found 369.2093.
2-Methyl-1-propyl-3-(7-ethyl-1-naphthoyl) indole, JWH-236. JWH-236 was prepared from 7-ethyl-1-naphthoic acid and 2-methyl-1-propylindole by method B. From 0.150 g (0.75 mmol) of 7-ethyl-1-naphthoic acid and 0.087 g (0.5 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 95:5) 0.163 g (92%) of JWH-236 as a pale brown oil: 1H NMR (300 MHz, CDCl3) δ 0.98 (t, J=7.3 Hz, 3H), 1.21 (t, J=7.6 Hz, 3H), 1.76-1.88 (m, 2H), 2.47 (s, 3H), 2.72 (q, J=7.6 Hz, 2H), 4.08 (t, J=7.4 Hz, 2H), 7.00 (t, J=7.3 Hz, 1H), 7.17 (t, J=7.3 Hz, 1H), 7.24 (d, J=7.6 Hz, 1H), 7.31 (d, J=8.2 Hz, 1H), 7.36-7.51 (m, 2H), 7.52 (d, J=6.0 Hz, 1H), 7.83 (d, J=8.2 Hz, 1H), 7.92 (d, J=8.2 Hz, 1H), 7.96 (s, 1H); 3C NMR (75.5 MHz, CDCl3) 811.4, 12.5, 15.5, 22.9, 29.2, 44.8, 109.4, 114.9, 121.3, 121.8, 122.1, 123.4, 124.0, 126.0, 127.1, 127.3, 128.1, 129.8, 130.6, 132.3, 136.1, 139.7, 142.9, 145.5, 193.6; MS (EI) m/z 355 (100), 338 (40); HRMS Calcd for C25H25NO: 355.1936, Found 355.1936.
2-Methyl-1-pentyl-3-(7-ethyl-1-naphthoyl)indole, JWH-262. JWH-262 was prepared from 7-ethyl-1-naphthoic acid and 2-methyl-1-pentylindole by method B. From 0.150 g (0.75 mmol) of 7-ethyl-1-naphthoic acid and 0.100 g (0.5 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 95:5) 0.167 g (87%) of JWH-262 as a pale brown oil: 1H NMR (300 MHz, CDCl3) δ 0.89 (t, J=6.9 Hz, 3H), 1.21 (t, J=7.6 Hz, 3H),1.26-1.43 (m, 4H), 1.74-1.79 (m, 2H), 2.47 (s, 3H), 2.71 (q, J=7.6 Hz, 2H), 4.09 (t, J=7.5 Hz, 2H), 6.99 (t, J=7.3 Hz, 1H), 7.16 (t, J=7.3 Hz, 1H), 7.24 (d, J=8.0 Hz, 1H), 7.30 (d, J=8.0 Hz, 1H), 7.36-7.48 (m, 2H), 7.52 (dd, J=7.1, 1.2 Hz, 1H), 7.83 (d, J=8.4 Hz, 1H), 7.91 (d, J=8.4 Hz, 1H), 7.97 (s, 1H); 13C NMR (75.5 MHz, CDCl3) δ 12.5, 13.9, 15.5, 22.3, 29.0, 29.3, 43.3, 53.3, 109.3, 114.9, 121.1, 121.7, 122.1, 123.4, 124.0, 126.0, 127.1, 127.2, 128.1, 129.7, 130.5, 132.5, 135.9, 139.6, 142.9, 145.4, 193.6; MS (EI) m/z 383 (100), 368 (50), 297 (50); HRMS Calcd for C27H29NO: 382.2175, Found 382.2175.
1-Propyl-3-(4-ethoxy-1-naphthoyl)indole, JWH-259. JWH-259 was prepared from 4-ethoxy-1-naphthoic acid and 1-propylindole by method B. From 0.13 g (0.6 mmol) of 4-ethoxy-1-naphthoic acid and 0.080 g (0.5 mmol) of 1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 9:1) 0.15 g (86%) of JWH-259 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.85 (t, J=7.3 Hz, 3H), 1.55 (t, J=6.9 Hz, 3H), 1.75-1.82 (m, 2H), 3.97 (t, J=7.1 Hz, 2H), 4.20 (q, J=6.9 Hz, 2H), 6.74 (d, J=7.9 Hz, 1H), 7.29-7.37 (m, 4H), 7.45-7.47 (m, 2H), 7.60 (d, J=7.9 Hz, 1H), 8.29-8.32 (m, 1H), 8.34-8.38 (m, 1H), 8.46-8.48 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.2, 14.7, 23.0, 48.5, 63.8, 102.7, 109.9, 117.6, 122.1, 122.5, 122.7, 123.3, 125.4, 125.5, 125.6, 125.7, 127.2, 127.9, 131.1, 132.1, 136.9, 137.4, 156.3, 191.7; HRMS Calcd for C24H23NO2: 357.1725, Found 357.1728.
1-Pentyl-3-(4-ethoxy-1-naphthoyl)indole, JWH-258. JWH-258 was prepared from 4-ethoxy-1-naphthoic acid and 1-pentylindole by method B. From 0.13 g (0.6 mmol) of 4-ethoxy-1-naphthoic acid and 0.094 g (0.5 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 9:1) 0.17 g (90%) of JWH-258 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.83 (t, J=6.6 Hz, 3H), 1.26-1.29 (m, 4H), 1.56 (t, J=6.9 Hz, 3H), 1.75-1.82 (m, 2H), 4.03 (t, J=7.1 Hz, 2H), 4.22 (q, J=6.9 Hz, 2H), 6.77 (d, J=7.9 Hz, 1H), 7.22-7.38 (m, 4H), 7.47-7.50 (m, 2H), 7.62 (d, J=7.9 Hz, 1H), 8.29-8.33 (m, 1H), 8.35-8.39 (m, 1H), 8.46-8.48 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 13.8, 14.7, 22.1, 28.8, 29.4, 46.9, 63.9, 102.8, 109.9, 117.6, 122.1, 122.5, 122.8, 123.3, 125.4, 125.5, 125.7, 125.8, 127.2, 127.9, 131.1, 132.1, 136.9, 137.3, 156.3, 191.7; HRMS Calcd for C26H27NO2: 385.2038, Found 385.2042.
2-Methyl-1-propyl-3-(4-ethoxy-1-naphthoyl)indole, JWH-261. JWH-261 was prepared from 4-ethoxy-1-naphthoic acid and 2-methyl-1-propylindole by method B. From 0.13 g (0.6 mmol) of 4-ethoxy-1-naphthoic acid and 0.087 g (0.5 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 9:1) 0.17 g (90%) of JWH-261 as a gum: 1H NMR (300 MHz, CDCl3) δ1.04 (t, J=7.2 Hz, 3H), 1.64 (t, J=6.8 Hz, 3H), 1.84-1.91 (m, 2H), 2.54 (s, 3H), 4.11 (t,J=7.2 Hz, 2H), 4.29 (q,J=6.8 Hz, 2H), 6.82 (d,J=7.9 Hz, 1H), 7.10 (t, J=7.5 Hz, 1H), 7.24 (t, J=7.2 Hz, 1H), 7.30-7.44 (m, 2H), 7.55-7.58 (m, 2H), 7.65 (d, J=7.9 Hz, 1H), 8.38-8.41 (m, 1H), 8.47-8.50 (m, 1H); 13C NMR (75.5 MHz, CDCl3) 611.3, 12.3, 14.6,22.8,44.6,63.6, 103.2, 109.3, 115.1, 121.1, 121.4, 121.8, 122.1, 125.3, 125.4, 125.7, 127.1, 127.3, 128.4, 131.8, 132.0, 135.9, 144.6, 156.4, 193.0; HRMS Calcd for C25H25NO2: 368.3411, Found 368.3403.
2-Methyl-1-pentyl-3-(4-ethoxy-1-naphthoyl)indole, JWH-260. JWH-260 was prepared from 4-ethoxy-1-naphthoic acid and 2-methyl-1-pentylindole by method B. From 0.13 g (0.6 mmol) of 4-ethoxy-1-naphthoic acid and 0.10 g (0.5 mmol) of 2-methyl-1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 9:1) 0.18 g (90%) of JWH-260 as a gum: 1H NMR (300 MHz, CDCl3) δ 0.89 (t, J=6.9 Hz, 3H), 1.34-1.36 (m, 4H), 1.56 (t, J=6.9 Hz, 3H), 1.74-1.81 (m, 2H), 2.47 (s, 3H), 4.07 (t, J=7.4 Hz, 2H), 4.22 (q, J=6.9 Hz, 2H), 6.75 (d, J=7.9 Hz, 1H), 6.99 (t, J=7.5 Hz, 1H), 7.15 (t, J=8.1 Hz, 1H), 7.22-7.30 (m, 2H), 7.45-7.50 (m, 2H), 7.55 (d, J=7.9 Hz, 1H), 8.26-8.29 (m, 1H), 8.36-8.39 (m, 1H); 13C NMR (75.5 MHz, CDCl3) 812.4, 13.9, 14.7, 22.3, 29.0, 29.3, 43.2, 63.8, 103.2, 109.3, 112.2, 115.2, 121.4, 121.9, 122.1, 125.3, 125.5, 125.7, 127.2, 127.3, 128.5, 131.8, 132.1, 135.9, 144.6, 156.5, 193.1; HRMS Calcd for C27H29NO2: 399.2193, Found 399.2198.
1-Propyl-3-(2-methoxy-1-naphthoyl)indole, JWH-265 (30). Indole 30 was prepared from 2-methoxy-1-naphthoic acid and 1-propylindole by method B. From 0.150 g (0.75 mmol) of 2-methoxy-1-naphthoic acid and 0.075 g (0.47 mmol) of 1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 7:3) 0.113 g (70%) of 1-propyl-3-(2-methoxy-1-naphthoyl)indole (30) as white crystals: m.p. 117-119° C.; 1H NMR (300 MHz, CDCl3) δ 0.77 (t, J=7.4 Hz, 3H), 1.70 (sept, J=7.2 Hz, 2H), 3.79 (s, 3H), 3.86 (t, J=6.8 Hz, 2H), 7.21-7.31 (m, 7H), 7.68-7.71 (m, 1H), 7.75-7.78 (m, 1H), 7.87 (d, J=9.0 Hz, 1H), 8.44 (s, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.0, 22.8, 48.4, 56.5, 109.9, 113.4, 117.9, 122.5, 122.6, 123.2, 123.7, 124.5, 125.1, 126.3, 126.8, 127.7, 128.6, 130.2, 131.8, 136.9, 137.9, 153.3, 190.7; MS (EI) m/z 343 (100), 326 (50), 172 (43); HRMS Calcd for C23H21NO2: 343.1572, Found 343.1577.
1-Pentyl-3-(2-methoxy-1-naphthoyl)indole, JWH-267 (32). Indole 32 was prepared from 2-methoxy-1-naphthoic acid and 1-pentylindole by method B. From 0.150 (0.75 mmol) of 2-methoxy-1-naphthoic acid and 0.090 g (0.48 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 7:3) 0.107 g (60%) of 32 as a viscous yellow gum: 1H NMR (300 MHz, CDCl3) δ 0.81 (t, J=6.9 Hz, 3H), 1.18-1.26 (m, 4H), 1.75 (quintet, J 2 032 7.1 Hz, 2H), 3.86 (s, 3H), 3.99 (t, J=7.2 Hz, 2H), 7.24-7.37 (m,7H), 7.68-7.72 (m,1H), 7.80-7.83 (m, 1H), 7.92 (d, J=9.1 Hz, 1H), 8.41 (s, 1H); 13C NMR (75.5 MHz, CDCl3) δ 22.0, 28.7, 29.2, 47.0, 56.5, 109.9, 113.4, 118.0, 122.6, 122.7, 123.2, 123.8, 124.6, 125.2, 126.4, 126.8, 127.7, 128.6, 130.2, 131.8, 136.9, 137.9, 153.4, 190.8; MS (EI) m/z 37 (100), 354 (61), 200 (55); HRMS Calcd for C25H25NO2: 371.1885, Found 371.1876.
2-Methyl-1-propyl-3-(2-methoxy-1-naphthoyl)indole, JWH-266 (31). Indole 31 was prepared from 2-methoxy-1-naphthoic acid and 2-methyl-1-propylindole by method B. From 0.150 g (0.75 mmol) of 2-methoxy-1-naphthoic acid and 0.086 g (0.50 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 7:3) 0.100 g (58%) of 31 as a yellow solid: m.p. 124-125° C.; 1H NMR (300 MHz, CDCl3) δ 0.86 (t, J=7.3 Hz, 3H), 1.57-1.73 (m, 2H), 3.73 (s, 3H), 3.92 (t, J=6.7 Hz, 2H), 7.08 (s, 3H), 7.17-7.30 (m, 7H), 7.58-7.60 (m, 1H), 7.74-7.76 (m, 1H), 7.87 (d, J=9.1 Hz, 1H); 1H NMR (500 MHz, DMSO-d6, 25° C.) δ 0.88 (br s, 3H), 1.61-1.78 (m, 2H), 3.78 (s, 3H), 4.08-4.23 (m, 2H), 7.13 (br s, 1H), 7.24-7.64 (m, 6H), 7.93-7.98 (m, 1H), 8.10 (d, J=9.1 Hz, 1H); 1H NMR (500 MHz, DMSO-d6, 125° C.) δ 0.88 (t, 6.8 Hz, 3H), 1.71-1.80 (m, 2H), 2.36 (s, 3H), 3.80 (s, 3H), 4.17 (t, J=7.4 Hz, 2H), 6.95 (t, J=7.8 Hz, 1H); 7.14 (td, J=0.5, 8.2 Hz, 1H), 7.32-7.39 (m, 2H), 7.47-7.58 (m, 4H), 7.91-7.96 (m,1H), 8.06 (d, J=9.2 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.0, 22.8, 44.4, 56.4, 109.3, 115.2, 121.4, 121.9, 122.2, 123.7, 123.9, 125.2, 126.3, 126.8, 127.7, 128.7, 130.0, 131.0, 135.9, 145.7, 152.8, 191.4; MS (EI) m/z 357 (44), 326 (100); HRMS Calcd for C24H23NO2: 357.1729, Found 357.1734.
2-Methyl-1-pentyl-3-(2-methoxy-1-naphthoyl)indole, JWH-267 (33). Indole 33 was prepared from 2-methoxy-1-naphthoic acid and 2-methyl-1-pentylindole by method B. From 0.150 g (0.75 mmol) of 2-methoxy-1-naphthoic acid and 0.094 g (0.47 mmol) of 2-methyl-1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 7:3) 0.109 g (60%) of 33 as a yellow solid: m.p. 39-40° C.; 1H NMR (300 MHz, CDCl3) δ 0.81 (t, J=7.0 Hz, 3H), 1.35-1.36 (m, 4H), 1.59-1.63 (m, 2H), 3.83 (s, 3H), 4.08 (t, J=7.0 Hz, 2H), 7.15 (s, 3H), 7.25-7.38 (m, 7H), 7.64-7.67 (m,1H), 7.80-7.84 (m, 1H), 7.94 (d, J=9.0 Hz, 1H); 2-Methyl-1-pentyl-3-(2-methoxy-1-naphthoyl)indole at 25; 1H NMR (500 MHz, toluene-d8, 25° C.) 80.72 (t, J=7.4 Hz, 3H), 0.87-096 (m, 2H), 0.96-1.07 (m, 2H), 1.18-1.27 (m, 2H), 2.15-2.74 (br, 3H), 3.32 (s, 3H), 3.41 (t, J=6.85 Hz, 2H), 6.88-7.25 (m,7H), 7.60-7.64 (m, 1H), 7.65 (d, J=9.2 Hz, 1H), 7.97 (d, J=7.8 Hz, 1H); 1H NMR (500 MHz, toluene-d8, 100° C.) d 0.72 (t, J=7.4 Hz, 3H), 0.97-1.11 (m, 4H), 1.30-1.39 (m, 2H), 2.34 (s, 3H), 3.41 (s, 3H), 3.54 (t, J=8.7 Hz, 2H), 6.89 (br s, 1H), 6.95-7.12 (m, 6H), 7.60 (d, J=7.3 Hz, 1H), 7.64 (d, J=10.2 Hz, 1H), 7.90 (d, J=8.2 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 13.9, 22.3, 29.0, 29.2, 43.2, 56.7, 109.3, 113.6, 116.0, 122.1, 122.2, 122.4, 123.9, 124.2, 125.2, 126.3, 127.0, 127.8, 128.9, 130.2, 131.2, 138.6, 152.9, 191.3; MS (EI) m/z 385 (34), 354 (100), 185 (46); HRMS Calcd for C26H27NO2: 385.2042, Found 385.2040.
6-Methoxy-1-naphthoic acid (35). To a solution of 2.3 g (13.2 mmol) of 6-methoxy-1-naphthol 40,41 in 50 mL of pyridine was added dropwise at 0° C. 2.6 mL (15.5 mmol) of trifluoromethanesulfonic anhydride. The solution was warmed to ambient temperature, stirred for 18 h, quenched with water and extracted with ether. The combined ethereal extracts were washed with 10% aqueous HCl until the aqueous solution was acidic. The solution was dried (MgSO4) and the solvent was removed in vacuo. The residue was chromatographed (petroleum ether:ethyl acetate, 12:1) to give 3.4 g (84%) of triflate as a yellow oil which was used in the subsequent step without further purification: 1H NMR (300 MHz, CDCl3) δ 3.92 (s, 3H), 7.16 (d, J=2.4 Hz, 1H), 7.27 (dd, J=8.7, 2.4 Hz, 2H), 7.41 (t, J=8.1 Hz, 1H), 7.95 (d, J=9.0 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 55.4, 106.0 115.3, 116.6, 120.7, 120.9, 121.6, 122.4, 125.8, 127.2, 136.5, 145.8, 158.7; MS (EI) m/z 306 (25), 173 (55), 145 (100).
To a solution of 0.96 g (3.18 mmol) of 6-methoxy-1-naphthyl trifluoromethanesulfonate in 30 mL of DMF was added 0.034 g (0.15 mmol) of Pd(OAc)2, 0.040 g (0.010 mmol) of 1,3-bis(diphenylphosphino)propane and 1.20 g of triethylamine. The mixture was flushed with carbon monoxide for 25 min, 3.1 mL of 96% formic acid was added dropwise and the reaction was stirred at ambient temperature for 6 h under an atmosphere of CO, diluted with water and extracted with ethyl acetate. The organic extracts were washed with five portions of brine, followed by two portions of aqueous NaHCO3. The bicarbonate extracts were combined, cautiously neutralized with 10% aqueous HCl, and extracted with ether. The ethereal extracts were washed with water, dried (MgSO4) and the solvent was removed in vacuo to give 0.42 g (65%) of acid 35 as a yellow powder. Recrystallization from ethyl acetate:petroleum ether gave pale yellow needles: m.p. 185-186° (lit mp 180-180.5° C., ref. 36); 1H NMR (300 MHz, CDCl3) δ 3.89 (s, 3H), 7.29 (dd, J=6.9, 2.4 Hz, 1H), 7.41 (d, J=2.4 Hz, 1H), 7.53 (t, J=7.8 Hz, 1H), 7.98 (d, J=6.1 Hz, 1H), 8.05 (d, J=8.1 Hz, 1H), 8.79 (d, J=9.1 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 55.2, 106.7, 120.0, 125.4, 126.1, 127.1, 127.5, 127.6, 131.8, 135.2, 157.1, 168.8; MS (EI) m/z 202 (100), 159 (35), 109 (50).
1-Propyl-3-(6-methoxy-1-naphthoyl)indole, JWH-163 (Method D). To a solution of 0.18 g (0.90 mmol) of 6-methoxy-1-naphthoic acid (35) in 10 mL of dichloromethane was added dropwise 0.44 ml (5.0 mmol) of oxalyl chloride. The solution was stirred at room temperature for 1 h, then heated at reflux for 1 h. After cooling, the solvent and residual oxalyl chloride were removed in vacuo. The residue was dissolved in 5 mL of toluene and added to a solution of 0.20 g (1.3 mmol) of 1-propylindole in 4 mL of toluene at 0° C. To this solution was added dropwise 0.82 mL (1.5 mmol) of 1.8 M EtAlCl2. The mixture was allowed to warm to ambient temperature and stirred for 18 h. After quenching with water, the reaction mixture was extracted with ether. The ethereal extracts were washed with water, dried (MgSO4) and the solvent was removed in vacuo. The residue was chromatographed (petroleum ether:ethyl acetate, 8:1) to give 0.19 g of JWH-163 as a colorless solid: m.p. 139-140° C.; 1H NMR (300 MHz, CDCl3) δ 0.87 (t, J=7.3 Hz, 3H), 1.81 (sextet, J=7.3 Hz, 2H), 3.91 (s, 3H), 4.01 (t, J=7.2 Hz, 2H), 7.12 (dd, J=9.0, 2.5 Hz, 1H), 7.18 (d, J=2.5 Hz, 1H), 7.30-7.39 (m, 4H), 7.44-7.52 (m, 2H), 7.82 (dd, J=7.5, 1.6 Hz, 1H), 8.09 (d, J=9.0 Hz, 1H), 8.47-8.50 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.3,23.0,48.7,55.2, 106.0, 110.0, 117.4, 119.3, 122.7, 122.8, 123.5, 123.6, 125.1, 126.1, 126.9, 127.5, 128.7, 135.1, 137.0, 138.0, 139.0, 157.7, 192.1; Anal. Calcd for C23H21NO2: C, 80.44; H, 6.16; N, 4.08; Found: C, 80.30; H, 6.17; N, 3.99.
1-Pentyl-3-(6-methoxy-1-naphthoyl)indole, JWH-166. JWH-166 was prepared from 6-methoxy-1-naphthoic acid and 1-pentylindole by method D. From 0.19 g (0.9 mmol) of acid and 0.24 g (1.3 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 8:1) 0.29 g (81%) of JWH-166 as an oil: 1H NMR (300 MHz,CDCl3) δ 0.85 (t, J=6.9 Hz, 3H), 1.23-1.33 (m, 4H), 1.75-1.85 (m, 2H), 3.98 (s, 3H), 4.05 (t, J=7.3 Hz, 2H), 7.12 (dd, J=9.1, 2.5 Hz, 1H), 7.18 (d, J=2.5 Hz, 1H), 7.33-7.40 (m, 4H), 7.47-7.52 (m, 2H), 7.83 (dd, J=7.4, 1.9 Hz, 1H), 8.10 (d, J=9.1 Hz, 1H), 8.45-8.48 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 13.8, 22.1, 28.9, 29.4, 47.1, 55.3, 106.1, 110.0, 117.5, 119.4, 122.8, 122.9, 123.5, 123.7, 125.1, 126.2, 127.0, 127.6, 128.7, 135.1, 137.0, 137.8, 139.0, 157.7, 192.1; MS (EI) m/z 371 (90), 354 (75), 314 (65), 214 (100); HRMS Calcd for C25H25NO2: 371.1885, Found 371.1885.
2-Methyl-1-propyl-3-(6-methoxy-1-naphthoyl)indole, JWH-151 (42). Indole 42 was prepared from 6-methoxy-1-naphthoic acid and 2-methyl-1-propylindole by method D. From 0.19 g (0.9 mmol) of acid and 0.22 g (1.3 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 8:1) 0.19 g (56%) of 42 as an oil: 1H NMR (300 MHz, CDCl3) δ 0.99 (t, J=7.5 Hz, 3H), 1.79-1.86 (m, 2H), 2.47 (s, 3H), 3.92 (s, 3H), 4.09 (t, J=7.5 Hz, 2H), 6.99 (t, J=7.3 Hz, 1H), 7.09 (dd, J=9.0, 2.7 Hz, 1H), 7.14-7.20 (m, 3H), 7.30 (d, J=7.8 Hz, 1H), 7.40-7.49 (m 2), 7.85 (d, J=7.5 Hz, 1H), 8.00 (d, J=9.0 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) 611.4, 12.5,22.9,44.8, 55.2, 106.1, 109.4, 114.9, 119.5, 121.2, 121.8, 122.1, 123.4, 125.7, 127.1, 127.2, 128.7, 135.2, 136.1, 140.5, 145.5, 157.7, 193.5; MS (EI) m/z 357 (100), 340 (40), 207 (98); HRMS Calcd for C24H23NO2: 357.1729, Found 357.1721.
2-Methyl-1-pentyl-3-(6-methoxy-1-naphthoyl)indole, JWH-153. JWH-153 was prepared from 6-methoxy-1-naphthoic acid and 2-methyl-1-pentylindole by method D. From 0.20 g (1.0 mmol) of acid and 0.24 g (1.2 mmol) of 2-methyl-1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 12:1) 0.23 g (60%) of JWH-153: m.p. 114.5-116° C.; 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=6.9 Hz, 3H), 1.35-1.38 (m, 4H), 1.74-1.79 (m, 2H), 2.46 (s, 3H), 3.92 (s, 3H), 4.10 (t, J=7.6 Hz, 2H), 6.98 (t, J=6.6 Hz, 1H), 7.08 (dd, J=9.3, 2.7 Hz, 1H), 7.13-7.20 (m 3), 7.29 (d, J=8.4 Hz, 1H), 7.39-7.48 (m, 2H), 7.84 (d, J=7.8 Hz, 1H), 8.98 (d, J=9.0 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 12.4, 13.8, 22.3, 29.0, 29.3, 43.3, 55.2, 106.1, 109.4, 114.9, 119.5, 121.2, 121.8, 122.1, 123.4, 125.6, 125.7, 127.1, 127.2, 128.7, 135.1, 136.0, 140.4, 145.4, 157.7, 193.4; Anal. Calcd for C26H27NO2: C, 81.01; H, 7.06; N, 3.63; Found: C, 80.80; H, 7.18; N, 3.62.
7-Methoxy-1-naphthoic acid (36). The trifluoromethanesulfonate ester of 7-methoxy-1-naphthol was prepared by the procedure employed for the preparation of the triflate of 6-methoxy-1-naphthol. From 2.30 g (15.5 mmol) of 7-methoxy-1-naphthol (40) 40,41 there was obtained after chromatography (petroleum ether:ethyl acetate, 12:1) 3.40 g (84%) of triflate as a colorless oil: 1H NMR (300 MHz, CDCl3) δ 3.91 (s, 3H), 7.16 (d, J=2.4 Hz, 1H), 7.27 (dd, J=8.7, 2.4 Hz, 2H), 7.41 (t, J=8.1 Hz, 1H), 7.71 (d, J=8.2 Hz, 1H), 7.95 (d, J=9.0 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) 655.4, 106.0, 115.3, 116.6, 120.7, 120.9, 121.6, 122.4, 125.8, 127.2, 136.5, 145.8, 158.7; MS (EI) m/z 306 (25), 173 (55), 145 (100).
Acid 36 was prepared from 7-methoxy-1-naphthyl trifluoromethanesulfonate by the procedure used for the preparation of acid 35. From 0.96 g (3.18 mmol) of triflate there was obtained, after recrystallization from ethyl acetate:petroleum ether, 0.35 g (54%) of acid 36: m.p. 185-186° (lit mp 169-170° C. ref. 37); 1H NMR (300 MHz, CDCl3) δ 3.89 (s, 3H), 7.29 (dd, J=6.9, 2.4 Hz, 1H), 7.41 (d, J=2.4 Hz, 1H), 7.53 (t, J=7.8 Hz, 1H), 7.98 (d, J=6.1 Hz, 1H), 8.05 (d, J=8.1 Hz, 1H), 8.79 (d, J=9.1 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 55.2, 106.7, 120.0, 125.4, 126.1, 127.1, 127.5, 127.6, 131.8, 135.2, 157.1, 168.8; MS (EI) m/z 202 (100), 159 (35), 109 (50).
1-Propyl-3-(7-methoxy-1-naphthoyl)indole JWH-165. JWH-165 was prepared from 7-methoxy-1-naphthoic acid and 1-propylindole by method D. From 0.18 g (0.9 mmol) of acid and 0.24 g (1.2 mmol) of 1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 8:1) 0.19 g (64%) of JWH-165: 1H NMR (300 MHz, CDCl3) δ 0.90 (t, J=8.1 Hz, 3H), 1.85 (q, J=7.2 Hz, 2H), 3.82 (s, 3H), 4.05 (t, J=7.2 Hz, 2H), 7.17 (dd, J=8.9, 2.4 Hz, 1H), 7.24-7.41 (m, 5H), 7.61-7.65 (m, 2H), 7.78 (d, J=8.9 Hz, 1H), 7.88 (d, J=8.1 Hz, 1H), 8.47-8.50 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 11.3, 23.1, 48.7, 55.3, 103.9, 110.0, 117.5, 119.4, 122.1, 122.7, 122.9, 123.5, 127.1, 129.4, 129.6, 130.0, 132.1, 137.0, 137.4, 137.8, 158.4, 192.2; MS (EI) m/z 343 (100), 326 (60), 314 (55), 207 (85); HRMS Calcd for C23H21NO2: 343.1572, Found 343.1581.
1-Pentyl-3-(7-methoxy-1-naphthoyl)indole, JWH-164. JWH-164 was prepared from 7-methoxy-1-naphthoic acid and 1-pentylindole by method D. From 0.19 g (1.0 mmol) of acid and 0.24 g (1.2 mmol) of 1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 8:1) 0.21 g (66%) of JWH-164: 1H NMR (300 MHz, CDCl3) δ 0.83 (t, J=7.5 Hz, 3H), 1.18-1.32 (m, 4H), 1.77-1.86 (m, 2H), 3.82 (s, 3H), 4.08 (t, J=7.2 Hz, 2H), 7.18 (dd, J=9.0, 2.5 Hz,1H), 7.34-7.42 (m, 5H), 7.61-7.65 (m, 2H), 7.79 (d, J=9.0 Hz, 1H), 7.89 (d, J=8.1 Hz, 1H), 8.46-8.51 (m, 1H); 13C NMR (75.5 MHz, CDCl3) δ 13.8, 22.1, 28.9, 29.5, 47.1, 55.3, 103.9, 110.0, 117.5, 119.4, 122.1, 122.2, 122.7, 122.9, 123.5, 127.1, 129.4, 129.6, 130.0, 132.1, 137.0, 137.4, 137.7, 158.4, 192.2; MS (EI) m/z 371 (100), 314 (55), 214 (45), 207 (65); HRMS Calcd for C25H25NO2: 371.1885, Found 371.1893.
2-Methyl-1-propyl-3-(7-methoxy-1-naphthoyl)indole, JWH-160. JWH-160 was prepared from 7-methoxy-1-naphthoic acid and 2-methyl-1-propylindole by method D. From 0.18 g (0.9 mmol) of acid and 0.24 g (1.2 mmol) of 2-methyl-1-propylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 8:1) 0.20 g (66%) of JWH-160 as a viscous oil: 1H NMR (300 MHz, CDCl3) δ 0.99 (t, J=7.3 Hz, 3H), 1.77-1.90 (m, 2H), 2.50 (s, 3H), 3.79 (s, 3H), 4.10 (t, J=7.5 Hz, 2H), 7.03 (t, J=7.8 Hz, 1H), 7.16-7.24 (m, 3H), 7.32 (t, J=7.1 Hz, 2H), 7.54-7.60 (m, 2H), 7.80 (d, J=8.9 Hz, 1H), 7.90 (d, J=7.8 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) d 11.4, 12.5, 22.9, 44.8,55.2, 103.7, 109.4, 115.3, 119.3, 121.2, 121.7, 122.1, 122.6, 127.2, 127.4, 129.5, 129.7, 130.1, 131.7, 136.1, 138.5, 145.1, 158.5, 193.8; MS (EI) m/z 357 (100), 340 (40), 300 (15); HRMS Calcd for C24H23NO2: 357.1729, Found 357.1720.
2-Methyl-1-pentyl-3-(7-methoxy-1-naphthoyl)indole, JWH-159. JWH-159 was prepared from 7-methoxy-1-naphthoic acid and 2-methyl-1-pentylindole by method D. From 0.18 g (0.9 mmol) of acid and 0.24 g (1.2 mmol) of 2-methyl-1-pentylindole there was obtained after chromatography (petroleum ether:ethyl acetate, 8:1) 0.19 g (43%) of JWH-159 as a viscous oil: 1H NMR (300 MHz, CDCl3) δ 0.91 (6.9, 3H), 1.18-1.26 (m, 4H), 1.80 (t, J=7.3 Hz, 2H), 2.50 (s, 3H), 3.79 (s, 3H), 4.12 (t, J=7.5 Hz, 2H), 7.01 (t, J=7.8 Hz, 1H), 7.15-7.24 (m, 3H), 7.32 (t, J=7.8 Hz, 2H), 7.55-7.59 (m, 2H), 7.80 (d, J=8.9 Hz, 1H), 7.89 (d, J=7.9 Hz, 1H); 13C NMR (75.5 MHz, CDCl3) δ 12.5, 13.9,22.4,29.1,29.4, 43.4,55.3, 103.7, 109.4, 115.0, 119.3, 121.2, 121.7, 122.1, 122.6, 127.2, 127.4, 129.5, 129.7, 130.1, 131.7, 136.0, 138.5, 145.0, 158.5, 193.8; MS (EI) m/z 385 (45), 328 (82), 207 (100); HRMS Calcd for C26H27NO2: 385.2041, Found 385.2039.
Receptor Binding Assays.
1. CB1 Assay. [3H] CP-55,940 (KD=690 nM) binding to P2 membranes was conducted as described elsewhere,50 except whole brain (rather than cortex only) was used. Displacement curves were generated by incubating drugs with 1 nM of [3H] CP-55,940. The assays were performed in triplicate, and the results represent the combined data from three individual experiments.
2.CB2 Assay. Human embryonic kidney 293 cells were maintained in Dulbecco's modified Eagle's medium (DMEM) with 10% fetal clone II (HyClone, Logan Utah) and 5% CO2 at 37° C. in a Form a incubator. Cell lines were created by transfection of CB2 pcDNA3 into 293 cells by the Lipofectamine reagent (Life Technologies, Gaithersburg, Md.). The human CB2 cDNA was a gift. Stable transformants were selected in growth medium containing geneticin (1 mg/mL, reagent, Life Technologies, Gaithersburg, Md.). Colonies of about 500 cells were picked (about 2 weeks post transfection) and allowed to expand, then tested for expression of receptor mRNA by northern blot analysis. Cell lines containing moderate to high levels of receptor mRNA were tested for receptor binding properties. Transfected cell lines were maintained in DMEM with 10% fetal clone II plus 0.3-0.5 mg/mL geneticin and 5% CO2 at 37° C. in a Form a incubator.
The current assay is a modification of Compton et al.42 Cells were harvested in phosphate-buffered saline containing 1 mM EDTA and centrifuged at 500 g. The cell pellet was homogenized in 10 mL of solution A (50 mM Tris-HCl, 320 mM sucrose, 2 mM EDTA, 5 mM MgCl2, pH 7.4). The homogenate was centrifuged at 1,600×g (10 min), the supernatant saved, and the pellet washed three times in solution A with subsequent centrifugation. The combined supernatants were centrifuged at 100,000×g (60 min). The (P2 membrane) pellet was resuspended in 3 mL of buffer B (50 mM Tris-HCl, 1 mM EDTA, 3 mM MgCl2, pH 7.4) to yield a protein concentration of approximately 1 mg/mL. The tissue preparation was divided into equal aliquots, frozen on dry ice, and stored at −70° C. Binding was initiated by the addition of 40-50 μg membrane protein to silanized tubes containing [3H]CP-55,940 (102.9 Ci/mmol) and a sufficient volume of buffer C (50 mM Tris-HCl, 1 mM EDTA, 3 mM MgCl2, and 5 mg/mL fatty acid free BSA, pH 7.4) to bring the total volume to 0.5 mL. The addition of 1 μM unlabelled CP-55,940 was used to assess nonspecific binding. Following incubation (30° C. for 1 hour), binding was terminated by the addition of 2 mL of ice cold buffer D (50 mM Tris-HCl, pH 7.4, plus 1 mg/mL BSA) and rapid vacuum filtration through Whatman GF/C filters (pretreated with polyethyleneimine (0.1%) for at least 2 hours). Tubes were rinsed with 2 mL of ice cold buffer D, which was also filtered, and the filters subsequently rinsed twice with 4 mL of ice cold buffer D. Before radioactivity was quantitated by liquid scintillation spectrometry, filters were shaken for 1 hr in 5 mL of scintillation fluid.
CP-55,940 and all cannabinoid analogs were prepared by suspension in assay buffer from a 1 mg/mL ethanolic stock without evaporation of the ethanol (final concentration of no more than 0.4%). When anandamide was used as a displacing ligand, experiments were performed in the presence of phenylmethylsulfonyl fluoride (50 μM). Competition assays were conducted with 1 nM [3H]-55,940 or 1 nM [3H]SR141716A and 6 concentrations (0.1 nM to 10 μM displacing ligands). Displacement IC50 values were originally determined by unweighted least-squares linear regression of log concentration-percent displacement data and then converted to Ki values using the method of Cheng and Prusoff.51 [3S]GTPγS Binding Assays. Materials. All chemicals were from Sigma (St. Louis, Mo.) except the following: [35S]GTPγS (1250 Ci/mmol) was purchased from New England Nuclear Group (Boston, Mass.), GTPγS from Boehringer Mannheim (New York, N.Y.), and DMEM/F-12 from Fischer Scientific (Pittsburgh, Pa.). Whatman GF/B glass fiber filters were purchased from Fisher Scientific (Pittsburgh, Pa.).
Membrane Preparations. Chinese Hamster Ovary (CHO) cells stably expressing the human CB2 receptor (CB213 CHO) were cultured in a 50:50 mixture of DMEM and Ham F-12 supplemented with 100 U/ml penicillin, 100 U/ml streptomycin, 0.25 mg/ml G418, and 5% fetal calf serum. Cells were harvested by replacement of the media with cold phosphate-buffered saline containing 0.4% EDTA followed by agitation. Membranes were prepared by homogenization of cells in 50 mM Tris-,HCl, 3 mM MgCl2, 1 mM EGTA, pH 7.4, centrifugation at 50,000×g for 10 minutes at 4° C., and resuspension in the same buffer at 1.5 mg/ml. Membranes were stored at −80° C. until use.
[35S]GTPγS Binding. Prior to assays, samples were thawed on ice, centrifuged at 50,000×g for 10 minutes at 4° C., and resuspended in Assay Buffer (50 mM Tris-HCl (pH 7.4), 3 mM MgCl2, 0.2 mM EGTA, and 100 mM NaCl). Reactions containing 10 Bg of membrane protein were incubated for 1.5 hr at 30° C. in Assay Buffer containing 10 BM GDP, 0.1 nM [35S]GTPγS, 0.1% bovine serum albumin, and various concentrations of agonist. Nonspecific binding was determined in the presence of 20 BM unlabeled GTPγS. Reactions were terminated by rapid vacuum filtration through GF/B glass fiber filters, and radioactivity was measured by liquid scintillation spectrophotometry at 95% efficiency for 35S.
Data Analysis. All data are reported as the means ± SEM of at least three experiments, each performed in triplicate. Nonlinear regression analysis was conducted by iterative fitting using JMP (SAS for Macintosh). Nonspecific [35S]GTPγS binding was subtracted from all data. Basal [35S]GTPγS binding is defined as specific [35S]GTPγS binding in the absence of drug. Net-stimulated [35S]GTPγS binding is defined as [35S]GTPγS binding in the presence of drug minus basal. Percent stimulation is expressed as (net stimulated [35S]GTPγS binding/basal)×100%.
Molecular Modeling
1. Conformational Analysis. The structures of the N-propyl-C-2H series analogs: JWH-163 and JWH-180; the N-propyl-C-2 methyl series analogs: JWH-151 and JWH189; the N-pentyl-C-2H series analogs: JWH-166, JWH-182, JWH-267; and, the N-pentyl C-2-methyl series analogs: JWH-153, JWH-181 and JWH-268 were built in the Spartan molecular modeling program (V4.1.1; Wavefunction, Inc. Irvine, Calif.). Each structure was minimized using the AM1, semi-empirical method. For each minimized structure, AM1 conformational searches were then performed for rotation about the carbonyl to indole C-3 and carbonyl to naphthoyl C-1 bonds. Coordinate Drive. In order to describe the energy barrier for rotation about the carbonyl oxygen-carbonyl carbon-naphthalene C-1-naphthalene C-2 torsion angle in JWH-268 (33), an AM1 coordinate drive study was undertaken using Spartan. The torsion angle was driven from its global minimum energy value of 99.47° to −80.49° in 10° increments. The single point energies of the lowest energy and highest energy conformers identified during this drive were calculated at the HF 3-21G* level using Spartan.
2. CB1 Receptor Docking Studies. Amino Acid Numbering System. In the discussion of receptor residues that follows, the amino acid numbering scheme proposed by Ballesteros and Weinstein52 is used. In this numbering system, the most highly conserved residue in each transmembrane helix (TMH) is assigned a locant of 50. This number is preceded by the TMH number and may be followed in parentheses by the sequence number. All other residues in a TMH are numbered relative to this residue. In this numbering system, for example, the most highly conserved residue in TMH 2 of the CB1 receptor is D2.50(163). The residue that immediately precedes it is A2.49(162).
Ligand/CB1 R* Complex. Because agonists are thought to have higher affinity for the activated form of GPCRs,53 agonist ligands in the work reported here were docked in a model of the activated state (R*) of CB1. This R* CB1 model was created by modification of our rhodopsin-based model of the inactive (R) form of CB1 and guided by the biophysical literature on the R to R* transition. A complete discussion of the creation of both our CB1 R and R* models can be found in our published work.47,54
A recent CB1 mutation study has suggested that residues F3.36, W5.43 and W6.48 are part of the binding site of the aminoalkylindole, WIN55212-2.47 These results are consistent with the earlier work of Shire which showed that the TMH4-E2-TMH5 region of CB1 contains residues critical for the binding of SR141716A and WIN55,212-2.55 Because the indoles studied here are structurally related to the aminoalkylindoles, we chose to dock the indoles studied here in the same region of CB1 (i.e., the TMH3-4-5-6 region). Each ligand was docked in the CB1 TMH 3-4-5-6 region using interactive computer graphics. For all analogs studied, the lowest energy s-trans conformer of each ligand was used for docking studies.46 The energy of the CB1+R* TMH bundle/ligand complex was minimized using the AMBER* united atom force field in Macromodel 6.5 (Schrodinger Inc., Portland, Oreg.). A distance dependent dielectric, 8.0 Å extended non-bonded cutoff (updated every 10 steps), 20.0 Å electrostatic cutoff, and 4.0 Å hydrogen bond cutoff were used. The first stage of the calculation consisted of 2000 steps of Polak-Ribier conjugate gradient (CG) minimization in which a force constant of 225 kJ/mol was used on the helix backbone atoms in order to hold the helix backbones fixed, while permitting the side chains to relax. The second stage of the calculation consisted of 100 steps of CG in which the force constant on the helix backbone atoms was reduced to 50 kJ/mol in order to allow the helix backbones to adjust. Stages one and two were repeated with the number of CG steps in stage two incremented from 100 to 500 steps until a gradient of 0.001 kJ/(mole Å2) was reached. Explicit hydrogens were included on all aromatic amino acid residues in order to better simulate aromatic stacking interactions.56
Each resultant receptor/ligand complex was analyzed for the presence of hydrogen bonding and aromatic stacking interactions. Aromatic stacking interactions were identified using Burley and Petsko's criteria. These investigators reported that aromatic-aromatic stacking interactions in proteins operate at distances (d) of 4.5 to 7.0 Å between ring centroids. The angle (a) between normal vectors of interacting aromatic rings typically is between 30° and 90°, producing a “tilted-T” or “edge-to-face” arrangement of interacting rings. Residues and/or ligand regions were designated here as participating in an aromatic stacking interaction if they had centroid to centroid distances between 4.5 Å and 7.0 Å. These interactions were further classified as “tilted-T” arrangements if 30°≦α≦90° and as parallel arrangements for a <30°. In interactions where a=0°, arrangements were also identified as offset or not offset, as Hunter and co-workers have shown that off-set parallel stacks are more energetically favorable.57
(10) Herkenham, M.; Lynn, A. B.; Little, M. D.; Johnson, M. R.; Melvin, L. S.; De Costa, D. R.; Rice, K. C. Proc. Natl. Acad. Sci. USA, 1990, 87, 1932.
While the invention has been described in terms of its preferred embodiments, those skilled in the art will recognize that the invention can be practiced with modification within the spirit and scope of the appended claims. Accordingly, the present invention should not be limited to the embodiments as described above, but should further include all modifications and equivalents thereof within the spirit and scope of the description provided herein.
This invention was made using funds from grants from the National Institutes of Health having grant numbers DA 03672 and DA 03590. The United States government may have certain rights in this invention.
| Number | Date | Country | |
|---|---|---|---|
| 60555355 | Mar 2004 | US | |
| 60477044 | Jun 2003 | US |
