David W. Pate

Cannabinoids in the treatment of glaucoma

The leading cause of irreversible blindness is glaucoma, a disease normally characterized by the development of ocular hypertension and consequent damage to the optic nerve at its point of retinal attachment. This results in a narrowing of the visual field, and eventually results in blindness. A number of drugs are available to lower intraocular pressure (IOP), but, occasionally, they are ineffective or have intolerable side-effects for some patients and can lose efficacy with chronic administration. The smoking of marijuana has decreased IOP in glaucoma patients. Cannabinoid drugs, therefore, are thought to have significant potential for pharmaceutical development. However, as the mechanism surrounding their effect on IOP initially was thought to involve the CNS, issues of psychoactivity hindered progress. The discovery of ocular cannabinoid receptors implied an explanation for the induction of hypotension by topical cannabinoid applications, and has stimulated a new phase of ophthalmic cannabinoid research. Featured within these investigations is the possibility that at least some cannabinoids may ameliorate optic neuronal damage through suppression of N-methyl-D-aspartate receptor hyperexcitability, stimulation of neural microcirculation, and the suppression of both apoptosis and damaging free radical reactions, among other mechanisms. Separation of therapeutic actions from side-effects now seems possible through a diverse array of novel chemical, pharmacological, and formulation strategies.

Ophthalmic arachidonylethanolamide decreases intraocular pressure in normotensive rabbits

Arachidonylethanolamide (AEA) was the first anandamide to be identified as an endogenous ligand for the cannabinoid receptor of porcine brain. Since cannabinoids have shown some value in the reduction of ocular hypertension, the title compound was evaluated in normotensive rabbits as a possible topically applied agent for reducing intraocular pressure. AEA was dissolved in an aqueous solution of 2-hydroxy-propyl-beta-cyclodextrin. Single eyedrops (25 microliters) containing 3.13, 6.25, 31.25, 62.5 or 125.0 micrograms of AEA were instilled unilaterally into eyes of normotensive albino and pigmented rabbits. The intraocular pressures (IOPs) of these rabbits were then measured at fixed time intervals. The effect of AEA on IOP in treated and untreated (contralateral) eyes was similar in both types of rabbits. Administration of 31.25 micrograms of AEA caused an immediate IOP reduction in the treated eyes. AEA doses of 62.5 micrograms caused an initial increase and subsequent decrease of IOP in the treated eyes. In the untreated eyes, a marginal ocular hypotensive response of limited duration occurred immediately after administration of AEA at doses 31.25 or 62.5 micrograms. A significant increase (without subsequent decrease below baseline) in IOP occurred in treated eyes after a dose of 125.0 micrograms. The lowest dose (3.13 micrograms) did not have an effect on IOP. This study constitutes the first published demonstration that topical, unilateral administration of AEA significantly decreases IOP in normotensive albino and pigmented rabbits. Although the mechanism of action by which this compound produces its hypotensive effect in the eye is not known, the results suggest that AEA may prove useful in the investigation of glaucoma therapy.

Effects of topical anandamides on intraocular pressure in normotensive rabbits

A series of anandamide-type compounds were synthesized and studied for their effect on the intraocular pressure (IOP) of normotensive pigmented rabbits. Each test compound was dissolved in an aqueous 2-hydroxypropyl-beta-cyclodextrin solution and administered (31.25 - 62.5 micrograms) unilaterally to the eye. The most promising anandamides caused a statistically significant reduction of IOP in treated eyes, usually preceded by an initial transient elevation of IOP, compared to saline controls. In the contralateral untreated eyes, only a marginal or short hypotensive response was observed. Indomethacin pre-treatment (12.5 mg, s.c.) eliminated the IOP response to administered anandamides and arachidonic acid.

Increase in aqueous solubility, stability and in vitro corneal permeability of anandamide by hydroxypropyl-β-cyclodextrin

Abstract Arachidonylethanolamide (AEA), an endogenous ligand for the cannabinoid receptor, has a low aqueous solubility and an instability which hinder its use in aqueous formulations. In the present study, the effect of cyclodextrins (CDs) on the aqueous solubility, stability and in vitro corneal permeability of AEA was studied. The corneal penetration of AEA in HP-β-CD formulations was investigated in vitro by using isolated corneas of rabbits. The phase solubility diagram with HP-β-CD was classified as Ap-type and stability constants (K1:1 and K1:2) for 1:1 and 1:2 inclusion complexes were calculated to be 39 419 M −1 and 12 M −1, respectively. The phase solubility diagram of AEA with DIME-β-CD and HP-γ-CD were of the A l -type, indicating the formation of 1:1-complexes. The stability constants for 1:1-complexes were 744 877 M−1 and 15 469 M−1, respectively. The complexation of AEA with HP-β-CD markedly increased the stability of AEA. The shelf-life (t90%) of AEA in 10.0% HP-β-CD solution at 50°C was determined to be 166 days. The complexation of AEA with HP-β-CD increased corneal penetration of AEA compared to a suspension of the compound. Maximum permeability was achieved with the lowest HP-β-CD concentration that dissolved AEA completely. The permeability of AEA correlated well with the concentration of free AEA in solution.

Hydroxypropyl-β-cyclodextrin and its combination with hydroxypropyl-methylcellulose increases aqueous solubility of Δ9-tetrahydrocannabinol

Abstract Δ9-tetrahydrocannabinol (THC) is the main psychoactive constituent of Cannabis sativa L. and its therapeutic effects are currently under intensive study. However, THC has a very low aqueous solubility (1–2 ( μg mL ), which restricts its use as a pharmaceutical. The present study demonstrates that THC forms a drug-cyclodextrin complex in an aqueous solution of hydroxypropyl-β-cyclodextrin (HP-β-CD), resulting in a thousand-fold increase in THC solubility. This improvement in solubility can be further increased by adding 0.1% hydroxypropylmethylcellulose to the HP-β-CD solution. The present results suggest that the use of cyclodextrins might be a simple and useful method to overcome the poor water solubility of THC.

Hydroxypropyl-β-cyclodextrin increases aqueous solubility and stability of anandamide

Anandamide (arachidonylethanolamide; AEA) is an endogenous ligand for the cannabinoid receptor and its pharmacological effects are under intensive study. However, AEA has a low aqueous solubility and stability which may restrict its use and may eventually endanger the reliability of the obtained results. In the present study, it was found that AEA forms inclusion complexes with cyclodextrins (CDs), resulting in greater aqueous solubility and stability of AEA as AEA/CD complex. Aqueous solubility increased 1 000 to 30 000-fold, depending on the type of CD (10% solution) used. The half-life of AEA in aqueous hydroxypropyl-beta-cyclodextrin solution (10%) at 50 degree C was 2.9 years. These results suggest that CD-technology will be a very useful method to overcome the solubility and stability problems of AEA.

Effects of topical alpha-substituted anandamides on intraocular pressure in normotensive rabbits

AbstractPurpose. Anandamides have been observed to lower intraocular pressure in the rabbit eye, preceded by a period of hypertension. Amidases are thought to catabolize these compounds into their component parts, including arachidonic acid. Direct application of arachidonic acid has been observed to cause a marked rise of intraocular pressure. Thus, anandamide analogs resistant to catabolism were thought possibly devoid of this initial hypertension, and their effects on rabbit IOP investigated. Methods. A series of chiral alpha-substituted anandamides were synthesized and studied for their effect on the intraocular pressure (IOP) of normotensive pigmented rabbits. Each test compound was dissolved in an aqueous 2-hydroxypropyl-β-cyclodextrin solution (containing 3% polyvinyl alcohol) and administered (62.5 μg) unilaterally to the eye. Results. The most promising compounds caused a statistically significant reduction of IOP (vs. vehicle) in the treated eyes. Of these, the R-alpha-isopropyl compound exhibited the best activity tested. Unlike the alpha-unsubstituted analogs previously studied, hypotensive effects were not preceded by an initial elevation of IOP and indomethacin pre-treatment (12.5 mg, s.c.) did not eliminate the IOP response, as demonstrated by administered R-alpha-isopropyl anandamide. Conclusions. Catabolism of alpha-unsubstituted anandamides may account for their observed intraocular hypertensive effects. The physiological mechanism by which alpha-substituted anandamides work apparently differs from that of the more easily metabolized alpha-unsubstituted compounds.

Effects of Topical Anandamide-Transport Inhibitors, AM404 and Olvanil, on Intraocular Pressure in Normotensive Rabbits

AbstractPurpose. To evaluate the effects of topically applied anandamide transport inhibitors, AM404 and olvanil, on the intraocular pressure (IOP) of normotensive rabbits. To determine if the ocular hypotension induced by topical anandamide (AEA) can be potentiated by co-administered AM404. Methods. Test compounds, in either hydroxypropyl-β-cyclodextrin (HP-β-CD) or propylene glycol, were administered unilaterally onto rabbit eyes. To determine if AM404 affects the IOP-profile of AEA, AM404 was administered ocularly 15 minutes before topical AEA. Phenylmethylsulfonyl fluoride (PMSF) (24 mg/kg, s.c.) was given 30 min before AEA to prevent its catabolism. IOPs of the treated and untreated eyes were measured. The cannabinoid agonist activities of AM404 and olvanil were studied by using [35S]GTPγS autoradiography. Results. Topical AM404 (62.5 μg), in HP-β-CD vehicle, decreased IOP significantly in treated eyes. AM404 (62.5 μg) induced a significant IOP increase without subsequent decrease when given in propylene glycol vehicle. Olvanil (312.5 μg) caused a significant IOP reduction without provoking an initial hypertensive phase. These compounds did not significantly affect the IOP of untreated eyes. Co-administered AM404 (125 μg in HP-β-CD) had no significant effect on the IOP profile of AEA (62.5 μg). Conclusions. Ocular administration of AM404 or olvanil decreased IOP in rabbits, although AM404 can provoke an initial ocular hypertension and did not potentiate the IOP responses induced by exogenous AEA.

Anandamide prodrugs. 1. Water-soluble phosphate esters of arachidonylethanolamide and R-methanandamide.

Phosphate esters of arachidonylethanolamide (AEA) and R-methanandamide were synthesized and evaluated as water-soluble prodrugs. Various physicochemical properties (pK(a), partition coefficient, aqueous solubility) were determined for the synthesized phosphate esters. The chemical stability of phosphate esters was determined at pH 7.4. In vitro enzymatic hydrolysis rates were determined in 10% liver homogenate, and in a pure enzyme-containing (alkaline phosphatase) solution at pH 7.4. The intraocular pressure (IOP) lowering properties of R-methanandamide phosphate ester were tested on normotensive rabbits. The phosphate promoiety increased the aqueous solubility of the parent compounds by more than 16500-fold at pH 7.4. Phosphate esters were stable in buffer solutions, but rapidly hydrolyzed to their parent compounds in alkaline phosphatase solution (t(1/2)<<15 s) and liver homogenate (t(1/2)=8-9 min). The phosphate ester of R-methanandamide reduced IOP in rabbits. These results indicate that the phosphate esters of AEA and R-methanandamide are useful water-soluble prodrugs.