fishman
Bluelighter
THE ENDOCANNABINOID SYSTEM:
Impediments and incentives towards its use as a novel target of future drug design
Although nearly thirty years of relative scientific stagnation passed between the initial isolations of delta-9-tetrahydrocannabinol (THC) from the cannabis plant in 1964 and anandamide from the human brain in 1992, cannabinoid receptors and their ligands have since been studied in depth over the past decade (primarily in Europe and Australia). There is no doubt that the results of these investigative efforts have been as intriguing as they have been therapeutically promising, yet there are still many more obstacles to be hurdled. While the implications of the American-led campaign against the use of cannabis on the acceptance and advancement of cannabinoid science could provide the foundation for an entire dissertation alone, these issues will not be significantly addressed in this paper. Rather, the specific obstacles of focus here are of a more pharmacologic nature. First, at the level of basic scientific investigation, experiments attempting to elucidate specific mechanisms of cannabinoid-related processes must become increasingly more robust in design. Results from such experiments should provide more compelling evidence that better demonstrates the [apparently] significant role that cannabinoids play in human physiology. Second, clinical investigations involving cannabinoid-related compounds must clearly prove to be at least as beneficial as currently available pharmaceuticals without exhibiting the undesirable memory-impairing effects of therapies relying solely on the exploitation of traditional, broader-acting CB1 / CB2 agonists like THC or anandamide. This can only be accomplished by better characterizing the subtle differences among receptors within the cannabinoid family (both isolated and putative), identifying the unique interactions between selective agonists / antagonists and such receptors so that potential therapeutic targets can be explored, and then rationally designing pharmaceutical solutions that a physician can utilize to better address the specific problem(s) of a patient in need.
The recent emergence of the endocannabinoid system as a seemingly dominant player in the overall regulation of general human physiology makes it imperative that all new experimental evidence be held up to closer scrutiny than in the past (Di Marzo, 2002; Pertwee, 2002). While the precise interactions between the endocannabinoid system and the other more well-characterized signal transduction systems within the body will most likely not be fully understood any time soon, the vast majority of recent scientific evidence points to the general conclusion that these interactions are quite significant, and that they are most likely much more complex than we may previously have thought (Di Marzo, 2002; Pertwee, 2002). Therefore, investigators are now allowed to assume less when analyzing certain types of experimental data because of the very real possibility that yet-to-be characterized interactions may be involved and are skewing their interpretation of the results. For example, it has been recently shown that anandamide is also an agonist of other specific receptor types that are structurally located entirely outside of the endocannabinoid system (CB1 / CB2), most notably the vanilloid receptor VR1. Anandamide (and related synthetic analogues) binds to VR1 receptors with significant affinity and efficacy, and the structural requirements for such binding are quite distinct from that of CB1 / CB2 receptors (Ralevic, 2001). Furthermore, these VR1 receptors are thought to be located both in the human brain and in many peripheral tissues, but the exact “downstream” signaling consequences of VR1 receptor activation by cannabinoids are not yet fully understood. In any event, the discovery and isolation of a structurally unique receptor family to which certain cannabinoid ligands can effectively bind forces investigators to return to the multitude of previous experiments involving CB1 and CB2 knockout mice and seriously reevaluate the old data in the context of this new knowledge. Assuming that knocking out CB1 and CB2 receptors was an accurate or equivalent representation of blocking anandamide action was obviously a flaw made in drawing past conclusions, as anandamide has since been clearly shown to continue to act both directly and indirectly on tissues lacking CB1, CB2, and VR1 receptors (Di Marzo, 2002). Hopefully, future investigators will assume less, and in doing so, will be able to more efficiently approach this challenging and uncharted territory.
Another example of an inherent stumbling block impeding the advancement of accurate scientific knowledge in this field is evidenced by a recent debate regarding the suspected presence of a specific transporter protein that was thought to be responsible for anandamide uptake through the post-synaptic membrane. For the past few years most researchers have agreed that experimental evidence suggested that some type of active transport mechanism was responsible for the “regulated” synaptic uptake that occurs prior to the hydrolysis of anandamide by fatty acid amide hydrolase (FAAH) once within the cell (Porter, 2001). This theoretical anandamide transporter became the subject of further investigations which attempted to inhibit this transport process via small-molecule blockade with the administration of novel anandamide-related compounds such as arvanil, olvanil, and N-(4-hydroxyphenyl)arachidonylamide (AM404). These compounds were all shown to significantly increase synaptic anandamide levels, and this was thought to occur due to a selective blockade of the anandamide transporter. Recently, however, it has been shown that all of these suspected “transporter blockers” were also inhibitors of FAAH in vitro. Unintentionally inhibiting FAAH activity would definitely decrease the rate of anandamide hydrolysis within the cell and most likely lead to the synaptic accumulation of anandamide observed in many earlier “transporter blocker” studies. Furthermore, when re-investigated on a shorter time scale (i.e., one minute or less) that is more appropriate for a rapidly-degraded neurotransmitter like anandamide, the suspected “transporter blockers” did not effectively increase synaptic anandamide levels in the same manner that the earlier transporter studies – those conducted on much longer time scales – had suggested (Glaser, 2003). It is now believed that anandamide uptake is entirely controlled by simple diffusion which itself is regulated by various metabolic factors (such as the rate of anandamide hydrolysis via intracellular FAAH) and “other downstream events.” (Glaser, 2003) This sequence of events surrounding the search for the transporter protein represents a great deal of lost time, money, and effort on the part of the researchers who prematurely attempted to blockade the theoretical “anandamide transporter” target without first better understanding the role FAAH played in anandamide breakdown. On a somewhat related note, I believe it is worth mentioning here that the Pharmacology 160 textbook devotes a few paragraphs of the chapter on bipolar disorder to the suggested potential use of omega-3 fatty acids as an effective treatment. My personal knowledge of biochemistry is limited, but I suspect that it may be possible that omega-3 fatty acids partially inhibit the activity of FAAH, which might in turn raise levels of synaptic anandamide (or some other non-cannabinoid neurotransmitter, perhaps) just enough to successfully manage the symptoms of bipolar disorder.
Once a more accurate characterization of the both the interworkings of the endocannabinoid system itself and the overall role it plays in human physiology has been established, the second major obstacle – undesirable clinical side effects – remains to be pharmacologically overcome. The most direct route of endocannabinoid manipulation has come from the synthesis of novel and selective (CB1 vs. CB2) receptor agonist and antagonist drugs. Direct agonists have been synthesized that are much more potent than both THC and anandamide and can be much more receptor-selective than either of the two as well, but these novel compounds have thus far primarily been utilized as tools for basic scientific investigation and not for therapeutic use. While these selective agonists suggest possible future use in a wide variety of treatment areas ranging from chronic pain to appetite disorders, we are far from the point of having physicians perscribr these compounds to patients; Marinol and its derivatives have been deemed sufficient for the time being. And again, the problem of the association of excessive CB1 receptor agonism with memory impairment continues to arise, as well as the potential for abuse (such CB1 agonism is also associated with the conspicuous dopamine reward pathways of much interest to those studying addiction and alcoholism; also, CB1 agonist exposure has recently been linked to increased preference for ethanol in mice) (Kim, 2001; Wang, 2003). But also, already there is much proof that the administration of novel CB1 antagonists can block the impairment of short-term memory that is often associated with THC or anandamide dosing; these antagonists bind to CB1 receptors and apparently block receptor action (Mallet, 1998). While such a drug may seem very desirable for those wishing to minimize the residual after-effects of recreational cannabis use (to successfully study for and take an exam after smoking earlier in the week, or for Monday morning on the job, etc.), little is known about the long-term effects of such rapid and intentional CB1 receptor manipulation, so this type of psychotherapy should be approached with an appropriate level of skepticism. CB1 antagonists may be useful in treating drug addiction, perhaps, but again, the long-term effects have yet to be studied (and the broader short-term effects have also barely been studied yet, for that matter) (Wang, 2003). A more promising route of subtle endocannabinoid manipulation has come from the disruption of anandamide-related metabolic processes to indirectly modulate the level of pre-existing synaptic anandamide (or perhaps even other known endocannabinoid agonists such as 2-arachidonoylglycerol, 2-AG). Selective inhibitors of FAAH, mentioned earlier, have been shown to reduce anxiety by reducing the rate of anandamide hydrolysis, which may then reduce general rates of anandamide syanaptic uptake (Kathuria, 2003). The final result, however, is an increase in anandamide levels through the CNS, which perhaps may have associated with it the same undesirable side effects of chronic marijuana use. Furthermore, side-effects due to long-term, selective inhibition of FAAH has not been studied extensively in a clinical setting. But the clincal acceptance of such uses may at least be a step in the right direction, perhaps, as everyone can agree that more selective agents are still obviously required.
A more promising area of cannabinoid-related therapeutics is that of CB2 receptor modulation, again primarily through the use of specific agonists / antagonists. The CB2 receptor subtype is expressed much less in the CNS when compared to expression-levels of CB1 and is predominantly thought to be closely associated with the peripheral, immunosuppressive effects of general cannabinoid agonists (Porter, 2001; Di Marzo, 2002). CB2 receptor modulation has been implicated in prospective cancer chemotherapies that exploit an overexpression of the receptor subtype in certain tumor cells as well (McKallip, 2002). Noticeable, undesirable side effects are less common when CB1 receptor manipulation is avoided, but it must be noted that these CB1-related side effects are typically cognitive in nature; CB2 receptor manipulation may lead to latent, non-cognitive, yet still undesirable consequences that may be just as damaging but not as strikingly apparent.
In the end, the entire field of endocannabinoid research is ripe for study, and the mystery of the complex interplay surrounding this recently discovered system is being unraveled and elucidated at an increasing pace. Only once the receptor subtypes and various endocannabinoid ligands are better understood will the organic chemists be able to design more highly specific compounds that target various points along the endocannabinoid metabolic pathways. At the present time, this seems to be the most promising attempt at influencing endocannabinoid functioning to better the lives of patients without introducing additional negative side effects. The promise of a new breed of psychotherapeutic drugs – perhaps most promising for those exhibiting comorbid alcoholism / depression / anxiety disorders – will most likely provide the motivation and financial backing for further investigations into the endocannabinoid system mainly because of their high profitability, but the overall result will still be a significant and beneficial increase in the body of knowledge regarding a seemingly important [but previously neglected] area of human molecular biology.
References
Aceto, M., Scates, S., Razdan, R., & Martin, B. (1998). Anandamide, an endogenous cannabinoid, has a very low physical dependence potential. The Journal of Pharmacology and Experimental Therapeutics. 287; 598-605.
Di Marzo, V., De Petrocellis, L., Fezza, F., Ligresti, A., & Bisogno, T (2002).
Anandamide receptors. Prostaglandins, Leukotrienes, and Essential Fatty Acids 66; 377-391.
Glaser, S., Abumrad, N., Fatade, F., et al. (2003, Apr.). Evidence against the
presence of an anandamide transporter. PNAS. 100; 4269-4274.
Kathuria, S., Gaetani, S., Fegley, D., Valisño, V., et al. (2003, Jan.). Modulation of anxiety through blockade of anandamide hydrolysis. Nature Medicine. 9; 76-81.
Kim, D. & Thayer, S. (2001). Cannabinoids inhibit the formation of new
synapses between hippocampal neurons in culture. The Journal of
Neuroscience. 21 (RC146); 1-5.
Mallet, P. & Beninger, R. (1998). The cannabinoid CB1 receptor antagonist
SR141716A attenuates the memory impairment produced by delta-9-
tetrahydrocannabinol or anandamide. Psychopharmacology. 140; 11-19.
McKallip, R., Lombard, C., Fisher, M., et al. (2002). Targeting CB2 cannabinoid receptors as a novel therapy to treat malignant lymphoblastic disease. Blood. 100; 627-634.
Pertwee, R. G. & Ross, T.A (2002). Cannabinoid receptors and their ligands.
Prostaglandins, Leukotrienes, and Essential Fatty Acids 66; 100-121.
Porter, A. & Felder, C. (2001). The endocannabinoid nervous system: unique opportunities for therapeutic intervention. Pharmacology & Therapeutics. 90; 45-60.
Ralevic, V., Kendall, D., Jerman, J., et al. (2001). Cannabinoid activation of recombinant and endogenous vanilloid receptors. European Journal of Pharmacology. 424; 211-219.
Wang, L., Liu, J., Harvey-White, J., Zimmer, A., & Kunos, G. (2003, Feb.).
Endocannabinoid signaling via cannabinoid receptor 1 is involved in ethanol preference and its age-dependent decline in mice. PNAS. 100; 1393-1398.
Impediments and incentives towards its use as a novel target of future drug design
Although nearly thirty years of relative scientific stagnation passed between the initial isolations of delta-9-tetrahydrocannabinol (THC) from the cannabis plant in 1964 and anandamide from the human brain in 1992, cannabinoid receptors and their ligands have since been studied in depth over the past decade (primarily in Europe and Australia). There is no doubt that the results of these investigative efforts have been as intriguing as they have been therapeutically promising, yet there are still many more obstacles to be hurdled. While the implications of the American-led campaign against the use of cannabis on the acceptance and advancement of cannabinoid science could provide the foundation for an entire dissertation alone, these issues will not be significantly addressed in this paper. Rather, the specific obstacles of focus here are of a more pharmacologic nature. First, at the level of basic scientific investigation, experiments attempting to elucidate specific mechanisms of cannabinoid-related processes must become increasingly more robust in design. Results from such experiments should provide more compelling evidence that better demonstrates the [apparently] significant role that cannabinoids play in human physiology. Second, clinical investigations involving cannabinoid-related compounds must clearly prove to be at least as beneficial as currently available pharmaceuticals without exhibiting the undesirable memory-impairing effects of therapies relying solely on the exploitation of traditional, broader-acting CB1 / CB2 agonists like THC or anandamide. This can only be accomplished by better characterizing the subtle differences among receptors within the cannabinoid family (both isolated and putative), identifying the unique interactions between selective agonists / antagonists and such receptors so that potential therapeutic targets can be explored, and then rationally designing pharmaceutical solutions that a physician can utilize to better address the specific problem(s) of a patient in need.
The recent emergence of the endocannabinoid system as a seemingly dominant player in the overall regulation of general human physiology makes it imperative that all new experimental evidence be held up to closer scrutiny than in the past (Di Marzo, 2002; Pertwee, 2002). While the precise interactions between the endocannabinoid system and the other more well-characterized signal transduction systems within the body will most likely not be fully understood any time soon, the vast majority of recent scientific evidence points to the general conclusion that these interactions are quite significant, and that they are most likely much more complex than we may previously have thought (Di Marzo, 2002; Pertwee, 2002). Therefore, investigators are now allowed to assume less when analyzing certain types of experimental data because of the very real possibility that yet-to-be characterized interactions may be involved and are skewing their interpretation of the results. For example, it has been recently shown that anandamide is also an agonist of other specific receptor types that are structurally located entirely outside of the endocannabinoid system (CB1 / CB2), most notably the vanilloid receptor VR1. Anandamide (and related synthetic analogues) binds to VR1 receptors with significant affinity and efficacy, and the structural requirements for such binding are quite distinct from that of CB1 / CB2 receptors (Ralevic, 2001). Furthermore, these VR1 receptors are thought to be located both in the human brain and in many peripheral tissues, but the exact “downstream” signaling consequences of VR1 receptor activation by cannabinoids are not yet fully understood. In any event, the discovery and isolation of a structurally unique receptor family to which certain cannabinoid ligands can effectively bind forces investigators to return to the multitude of previous experiments involving CB1 and CB2 knockout mice and seriously reevaluate the old data in the context of this new knowledge. Assuming that knocking out CB1 and CB2 receptors was an accurate or equivalent representation of blocking anandamide action was obviously a flaw made in drawing past conclusions, as anandamide has since been clearly shown to continue to act both directly and indirectly on tissues lacking CB1, CB2, and VR1 receptors (Di Marzo, 2002). Hopefully, future investigators will assume less, and in doing so, will be able to more efficiently approach this challenging and uncharted territory.
Another example of an inherent stumbling block impeding the advancement of accurate scientific knowledge in this field is evidenced by a recent debate regarding the suspected presence of a specific transporter protein that was thought to be responsible for anandamide uptake through the post-synaptic membrane. For the past few years most researchers have agreed that experimental evidence suggested that some type of active transport mechanism was responsible for the “regulated” synaptic uptake that occurs prior to the hydrolysis of anandamide by fatty acid amide hydrolase (FAAH) once within the cell (Porter, 2001). This theoretical anandamide transporter became the subject of further investigations which attempted to inhibit this transport process via small-molecule blockade with the administration of novel anandamide-related compounds such as arvanil, olvanil, and N-(4-hydroxyphenyl)arachidonylamide (AM404). These compounds were all shown to significantly increase synaptic anandamide levels, and this was thought to occur due to a selective blockade of the anandamide transporter. Recently, however, it has been shown that all of these suspected “transporter blockers” were also inhibitors of FAAH in vitro. Unintentionally inhibiting FAAH activity would definitely decrease the rate of anandamide hydrolysis within the cell and most likely lead to the synaptic accumulation of anandamide observed in many earlier “transporter blocker” studies. Furthermore, when re-investigated on a shorter time scale (i.e., one minute or less) that is more appropriate for a rapidly-degraded neurotransmitter like anandamide, the suspected “transporter blockers” did not effectively increase synaptic anandamide levels in the same manner that the earlier transporter studies – those conducted on much longer time scales – had suggested (Glaser, 2003). It is now believed that anandamide uptake is entirely controlled by simple diffusion which itself is regulated by various metabolic factors (such as the rate of anandamide hydrolysis via intracellular FAAH) and “other downstream events.” (Glaser, 2003) This sequence of events surrounding the search for the transporter protein represents a great deal of lost time, money, and effort on the part of the researchers who prematurely attempted to blockade the theoretical “anandamide transporter” target without first better understanding the role FAAH played in anandamide breakdown. On a somewhat related note, I believe it is worth mentioning here that the Pharmacology 160 textbook devotes a few paragraphs of the chapter on bipolar disorder to the suggested potential use of omega-3 fatty acids as an effective treatment. My personal knowledge of biochemistry is limited, but I suspect that it may be possible that omega-3 fatty acids partially inhibit the activity of FAAH, which might in turn raise levels of synaptic anandamide (or some other non-cannabinoid neurotransmitter, perhaps) just enough to successfully manage the symptoms of bipolar disorder.
Once a more accurate characterization of the both the interworkings of the endocannabinoid system itself and the overall role it plays in human physiology has been established, the second major obstacle – undesirable clinical side effects – remains to be pharmacologically overcome. The most direct route of endocannabinoid manipulation has come from the synthesis of novel and selective (CB1 vs. CB2) receptor agonist and antagonist drugs. Direct agonists have been synthesized that are much more potent than both THC and anandamide and can be much more receptor-selective than either of the two as well, but these novel compounds have thus far primarily been utilized as tools for basic scientific investigation and not for therapeutic use. While these selective agonists suggest possible future use in a wide variety of treatment areas ranging from chronic pain to appetite disorders, we are far from the point of having physicians perscribr these compounds to patients; Marinol and its derivatives have been deemed sufficient for the time being. And again, the problem of the association of excessive CB1 receptor agonism with memory impairment continues to arise, as well as the potential for abuse (such CB1 agonism is also associated with the conspicuous dopamine reward pathways of much interest to those studying addiction and alcoholism; also, CB1 agonist exposure has recently been linked to increased preference for ethanol in mice) (Kim, 2001; Wang, 2003). But also, already there is much proof that the administration of novel CB1 antagonists can block the impairment of short-term memory that is often associated with THC or anandamide dosing; these antagonists bind to CB1 receptors and apparently block receptor action (Mallet, 1998). While such a drug may seem very desirable for those wishing to minimize the residual after-effects of recreational cannabis use (to successfully study for and take an exam after smoking earlier in the week, or for Monday morning on the job, etc.), little is known about the long-term effects of such rapid and intentional CB1 receptor manipulation, so this type of psychotherapy should be approached with an appropriate level of skepticism. CB1 antagonists may be useful in treating drug addiction, perhaps, but again, the long-term effects have yet to be studied (and the broader short-term effects have also barely been studied yet, for that matter) (Wang, 2003). A more promising route of subtle endocannabinoid manipulation has come from the disruption of anandamide-related metabolic processes to indirectly modulate the level of pre-existing synaptic anandamide (or perhaps even other known endocannabinoid agonists such as 2-arachidonoylglycerol, 2-AG). Selective inhibitors of FAAH, mentioned earlier, have been shown to reduce anxiety by reducing the rate of anandamide hydrolysis, which may then reduce general rates of anandamide syanaptic uptake (Kathuria, 2003). The final result, however, is an increase in anandamide levels through the CNS, which perhaps may have associated with it the same undesirable side effects of chronic marijuana use. Furthermore, side-effects due to long-term, selective inhibition of FAAH has not been studied extensively in a clinical setting. But the clincal acceptance of such uses may at least be a step in the right direction, perhaps, as everyone can agree that more selective agents are still obviously required.
A more promising area of cannabinoid-related therapeutics is that of CB2 receptor modulation, again primarily through the use of specific agonists / antagonists. The CB2 receptor subtype is expressed much less in the CNS when compared to expression-levels of CB1 and is predominantly thought to be closely associated with the peripheral, immunosuppressive effects of general cannabinoid agonists (Porter, 2001; Di Marzo, 2002). CB2 receptor modulation has been implicated in prospective cancer chemotherapies that exploit an overexpression of the receptor subtype in certain tumor cells as well (McKallip, 2002). Noticeable, undesirable side effects are less common when CB1 receptor manipulation is avoided, but it must be noted that these CB1-related side effects are typically cognitive in nature; CB2 receptor manipulation may lead to latent, non-cognitive, yet still undesirable consequences that may be just as damaging but not as strikingly apparent.
In the end, the entire field of endocannabinoid research is ripe for study, and the mystery of the complex interplay surrounding this recently discovered system is being unraveled and elucidated at an increasing pace. Only once the receptor subtypes and various endocannabinoid ligands are better understood will the organic chemists be able to design more highly specific compounds that target various points along the endocannabinoid metabolic pathways. At the present time, this seems to be the most promising attempt at influencing endocannabinoid functioning to better the lives of patients without introducing additional negative side effects. The promise of a new breed of psychotherapeutic drugs – perhaps most promising for those exhibiting comorbid alcoholism / depression / anxiety disorders – will most likely provide the motivation and financial backing for further investigations into the endocannabinoid system mainly because of their high profitability, but the overall result will still be a significant and beneficial increase in the body of knowledge regarding a seemingly important [but previously neglected] area of human molecular biology.
References
Aceto, M., Scates, S., Razdan, R., & Martin, B. (1998). Anandamide, an endogenous cannabinoid, has a very low physical dependence potential. The Journal of Pharmacology and Experimental Therapeutics. 287; 598-605.
Di Marzo, V., De Petrocellis, L., Fezza, F., Ligresti, A., & Bisogno, T (2002).
Anandamide receptors. Prostaglandins, Leukotrienes, and Essential Fatty Acids 66; 377-391.
Glaser, S., Abumrad, N., Fatade, F., et al. (2003, Apr.). Evidence against the
presence of an anandamide transporter. PNAS. 100; 4269-4274.
Kathuria, S., Gaetani, S., Fegley, D., Valisño, V., et al. (2003, Jan.). Modulation of anxiety through blockade of anandamide hydrolysis. Nature Medicine. 9; 76-81.
Kim, D. & Thayer, S. (2001). Cannabinoids inhibit the formation of new
synapses between hippocampal neurons in culture. The Journal of
Neuroscience. 21 (RC146); 1-5.
Mallet, P. & Beninger, R. (1998). The cannabinoid CB1 receptor antagonist
SR141716A attenuates the memory impairment produced by delta-9-
tetrahydrocannabinol or anandamide. Psychopharmacology. 140; 11-19.
McKallip, R., Lombard, C., Fisher, M., et al. (2002). Targeting CB2 cannabinoid receptors as a novel therapy to treat malignant lymphoblastic disease. Blood. 100; 627-634.
Pertwee, R. G. & Ross, T.A (2002). Cannabinoid receptors and their ligands.
Prostaglandins, Leukotrienes, and Essential Fatty Acids 66; 100-121.
Porter, A. & Felder, C. (2001). The endocannabinoid nervous system: unique opportunities for therapeutic intervention. Pharmacology & Therapeutics. 90; 45-60.
Ralevic, V., Kendall, D., Jerman, J., et al. (2001). Cannabinoid activation of recombinant and endogenous vanilloid receptors. European Journal of Pharmacology. 424; 211-219.
Wang, L., Liu, J., Harvey-White, J., Zimmer, A., & Kunos, G. (2003, Feb.).
Endocannabinoid signaling via cannabinoid receptor 1 is involved in ethanol preference and its age-dependent decline in mice. PNAS. 100; 1393-1398.
