36


Development of Cannabis-based therapeutics

David W. Pate

HortaPharm B.V., Amsterdam, The Netherlands


        Below appears the edited transcript of a lecture given by IHA Secretary David Pate at the National Academy of Sciences in Washington, D. C. on February 24, 1998, as part of the Institute of Medicine study to evaluate the therapeutic value of marijuana and its chemical components. (Editor's note: see page 53.)


Female Cannabis inflorescence

Female Cannabis inflorescence with THC-containing resin glands.

        MR. PATE: I work for a company that is, in the literal sense, unique in the world. HortaPharm is a company that was founded basically by Americans who thought it pointless to try to get the American government to approve what we were doing. So, we went to Holland and, with a lot of work, we finally got approval for growing Cannabis as a potential raw material for the manufacture of pharmaceuticals. You should know that we are not in the marijuana business per se. We see this as a means to an end.
        That is an example of our means, a picture of our greenhouse. We grow this material essentially to use as the raw material for the production of Δ9-tetrahydrocannabinol (THC), as I just mentioned, but you see those little white areas in the middle of the green leaves? Those are the floral clusters.
        I will give you a little bit of a closer look. That is a close-up example of the floral clusters. You can see that the little dots sprinkled over the surface, that give it an encrusted look, are the primary sites of cannabinoid manufacture on the plant. These are small glands that are, broadly speaking, maybe 50 to 150 microns in diameter, at least the reservoirs are. They usually sit on small multi-cellular pedestals. You can imagine a tiny golf ball and golf tee situated on the surface of the leaf. That is a reasonable analogy. THC stored in these reservoirs are quite stable if the flowers are handled carefully and the glands are left intact. This contrasts with the instability of chemically isolated THC, especially in solutions at room temperature, particularly when exposed to light.
        The synthesis of THC, as we heard, is an expensive process. We figured that direct agronomic production was probably less so. A parallel might be the production of morphine, which on an academic level, with modern techniques, is synthesizable from scratch, but no one does that. They extract poppies in order to produce morphine.
        We have produced plants that, at least for the last season, were about 16 percent THC. This is done with no extraordinary biotechnological efforts; in other words, no gene splicing or anything of that sort.
        This is a picture of the four candidate compounds that we are involved with. You hear a lot about 60-odd cannabinoids or 400-odd compounds in Cannabis. Basically, this claim is a summary of the collective records of all researchers over the past decades finding molecules in any detectable quantity in all the varieties of Cannabis investigated. In truth, or at least in perspective, Cannabis contains mostly these four compounds. These are Δ9-THC, cannabichromene, cannabidiol and cannabigerol. The biogenetic evolution of the cannabinoid compounds begins at cannabigerol, which is labeled in the slide as CBG and from that starting point comes cannabichromene (CBC), cannabidiol (CBD), and THC.
        Now, for a long time there was a theory that THC came from CBG via CBD. So CBG was the first biogenetic cannabinoid, CBC came from that, CBD came from that, and THC came from CBD. There is now direct evidence, via enzyme studies, that at least in some strains, the THC comes directly from CBG, bypassing the CBD step, if indeed the CBD theory is correct. That theory has not been unambiguously proven yet, in spite of its age. A few strains of Cannabis have absolutely none of the CBD, which led people initially to suspect that it came from some other source, probably directly from CBG rather than indirectly via CBD.
        There are a couple of other compounds that are found fairly frequently. That is cannabinol (CBN), which is a fully aromatic version of THC, and Δ8-THC, which has a double bond in the other direction. These are now seen as either degradation products or artifacts of the analysis. Δ8-THC is about two-thirds as potent as Δ9-THC, and cannabinol occurs fairly frequently in old Cannabis. In fresh Cannabis, it is not usually found.
        Each of the four main cannabinoids are present in varying amounts. CBG is normally present in the least amount. CBD, in temperate plants, plants growing in the midwest US or Europe, are high in this compound, but low in THC. Plants growing in the tropics usually have the inverse ratio. For a long time, CBC was hard to separate from CBD by gas chromatography (GC). So, the reportage of either of these compounds in the literature before the late 1970s is suspect. Tropical plants tend to have more CBC than CBD, and temperate plants the inverse ratio.
        Our basic premise, besides the fact that the synthesis is a bit complicated, was that perhaps we could have the plant, being the chemist that it is, manufacture a relatively clean profile in the resin it produces. So, a lot of downstream chromatographic processing simply isn't necessary because there isn't anything else there.
        This is an example of a nice clean GC profile. The peak on the left is an internal standard, and to the right is THC. You see a little bit of other peaks here and there, but nothing significant. We have been able, with a simple alcohol extraction, to get solutions that were, by our estimates, when shot into the GC directly after filtration, over the FDA 95% minimum specifications for Marinol. We've gotten 95 to 97 percent THC. The pharmacopoeia also specifies that Marinol can have up to two percent Δ8-THC as an impurity, and that much is never found here. So, loosely speaking, you might say that we could approach FDA specifications "on the hoof". This GC trace is not the best example we have, but it is one of the better ones.
        Well, what is this all used for? We have heard a lot about the indications. The first two shown in the THC slide each have an asterisk because these are the FDA approved uses. We have heard a lot of other possible uses today. Spasticity from spinal injury or MS, glaucoma is another possibility, pain, inflammation, insomnia, and asthma.
        The use of medical marijuana for asthma is somewhat at cross purposes, that is, the fact that people are inhaling smoke. So, it is a mixed bag of results, initial irritation and subsequent long-term help, "long-term" meaning quite a few hours.
        Something that isn't appreciated so much as a potential new drug is CBD. There is a lot of it in certain strains of Cannabis. We have some strains of Cannabis that look like the previous GC trace, only that large peak is CBD instead of THC. So, we can get extremely clean profiles of CBD from the plant, essentially single component mixtures. Developing an approved pharmaceutical from this compound is beyond our company's ability, for reasons that have been alluded to in the earlier talks. It costs a lot of money. But there is considerable potential for CBD. One of the more interesting potentials is in the treatment of schizophrenia. It also has some anxiolytic effects. There is only one group, in Brazil, that has looked deeply into the use of CBD for schizophrenia. Unfortunately, the amounts involved are reasonably large, as far as drug effects are involved, and that is on the order of a quarter gram or half a gram per dose. To treat a person in a long-term study, let alone a number of patients, requires quite a bulk of this compound. We have been in correspondence with these researchers and have received requests for supply. If we ever get the time to do so, we will provide them with the 200 or 300 grams of this material that they need to start their study.
        One other utility for this compound that I will mention, just sort-of for the novelty value, is that of food preservation. There have been a number of studies that have shown that spiking foods with CBD acid on the order of 10 parts per million helps to prevent spoilage. CBD acid has also been found to have antibiotic effects against gram-positive bacteria.
        It should be noted that the compounds that I displayed initially, the four compounds, normally are not present as just phenolic compounds in the plant. They have a carboxylic acid moiety attached. This is usually ortho to the phenolic group on the C ring, the right-hand-side ring. What happens, though, is it is easily decarboxylated with the application of heat. Anything over, say, 110 or 120 degrees Celsius will very rapidly cause that carboxylic group to disappear as water and CO 2 , although this also happens slowly at room temperature.
        The carboxylic acid species are not orally active in a significant sense. You have to heat them up. If you were going to take oral medical marijuana, you would have to cook it in order to decarboxylate these compounds, although a certain percentage is naturally decarboxylated by the fact of the plant staying outside in the sun all day. However, the percentages of that happening are actually fairly low. However, when you smoke the materials, the in situ heating of the pipe or the joint, or whatever it is, causes that decarboxylation to occur. This renders the materials not only more absorbable, but it helps to make them more volatile because the carboxylic acid groups of the native compounds have more hydrogen bonding.
        There are a couple of other cannabinoids to mention. CBC is sometimes found in substantial quantities, but often very low quantities. As I mentioned, its resolution by gas chromatography from CBD had been impossible, up until a comparatively few years ago. It's still not that easy. CBC has some -- Dr. ElSohly can probably address this better -- it has some effects on bacteria, as does CBG on fungi. CBC also has some anti-inflammatory effects.
        Basically, there are three medical objections to using the herbal form of Cannabis as a therapeutic agent and I will add maybe a fourth one. (It is sort-of an adjunct to objection number three).
        First of all, most natural products contain more than one thing. They contain a range of compounds. Actually, sometimes this can be advantageous, for example, providing synergisms. However, sometimes these other things might be toxic or act in unpredictable ways. In any case, modern medical science likes single component "silver bullets", rather than multi-component "herbal shot-guns".
        The next problem is that the chemical constituents vary. Most of this is under genetic control, which is a fortunate thing for us, but some of it has to do with where and how it is grown. The latitudes, the weather, the soil conditions, a lot of variables are potentially involved. With Cannabis, variability in chemical constituents seems to derive from its exposure to environmental stress. A stressed plant will often produce more of certain chemical constituents. For example, Cannabis exposed to ultraviolet radiation produces substantially more THC. This may have something to do with the fact that you have more potent Cannabis in the tropical regions. This is both from an environmental perspective in the immediate sense, and an evolutionary perspective in the sense of a long-term selection process that may have occurred.
        The third objection, of course, is pyrolysis products. Medical science doesn't like smoke in the lungs, and there are some reasons for that. How practical a problem this is depends primarily on longevity of the dosing. If you are talking about somebody going through six weeks of chemotherapy, or two or three six-week bouts, or less than a year of use, it is probably irrelevant. You are probably not going to have any real ill effects from that, especially compared with the horribly toxic anti-cancer drugs administered.
        If you are talking about somebody who has MS, who is 25 years old and who might, with luck, expect to live for another 25 or more years -- I don't actually know what the life expectancy is, it probably varies -- then you are talking about chronic dosing daily for quite a few years.
        Of course, the other variable is how much dosing is necessary. You take a disease like glaucoma, and you have somebody who is middle-aged, perhaps, smoking pretty-much like a chimney because the dose required is substantial and must be maintained all day and for a future life expectancy of decades. On the other hand, with the previous MS example, where the medical dose for THC is just at the borderline of psychoactivity and taken much less frequently, on demand, it is not really that much. Well, then you perhaps have less problem with a chronic dosing of this more limited amount of smoke.
        Compounding this is yet another dimension of consideration, the potency of the plant you are using. Research has shown that the more potent the marijuana, the less tars and carbon monoxide are inhaled per effective dose. Going from a 2% THC content to a 4% THC content material reduces by two thirds the tars and carbon monoxide absorbed by the subject per milligram of THC delivered. This fortunate trend probably holds or perhaps is even greater for the more potent varieties of marijuana available. It certainly makes sense that the fewer inhalations it requires to achieve the desired effect, the less smoke damage is possible.
        Adding to the third medical objection is a fourth possible objection, that of inhaling not only smoke, but potentially parasitic organisms. It doesn't seem that, on the whole, there has been a lot of problems with that in the general population. There are occasional citations in the medical literature, but considering the tens of millions of people who are involved with Cannabis on the recreational level in the US alone, it is apparently not too frequent. There is more concern about this possible problem for AIDS or cancer chemotherapy immunocom-promised patients. Problems like that I haven't heard of, so much either, but in theory you would be more cautious about something like that.
        Well, these are technical problems, and technical problems generally have technical answers. Again, our Cannabis has not necessarily been designed for direct use as an herbal medicament, although certainly it is amenable to that use, should the demand arise. So, what we have done is, through a careful process of selective breeding, we have gotten plants that essentially contain a single component, which by natural product standards, is somewhat extraordinary. Now, I am using the term single component in a very loose way. There are minor traces of other cannabinoids and there are tiny amounts of terpenes, which actually are the materials that you smell, since the cannabinoids don't have a native smell.
        The second technical answer we employ is the controlled production of clonal Cannabis. If you are careful and you grow it the same way, the same genetic stock will produce the same balance of cannabinoids time after time, season after season, eight or ten years so far, in our experience. Why is that? Because if you are growing clonal Cannabis, you are essentially growing the same plant. Basically, you take the plant, you make a twin, and you grow out one for your own purposes and you keep the other for library stock. If it is grown under reasonably similar conditions, even the quantitation is very similar. Certainly the qualitative profile is identical. We have variability in quantitation, but that can be taken into account by analysis: 12 percent, 14, 16 percent, etc.
        The third technical answer we have achieved is non-pyrolytic vaporization of cannabinoids. We have been working on the last two or three years on a vaporization device that supersedes the technology that you see in magazines and in "head shops". Recent results have demonstrated that "tars" are generated by burning cellulose, the "wood" of marijuana. The active ingredients are probably not responsible. In other words, completely extracted placebo Cannabis has been shown to generate as much tar as 4% THC content Cannabis. So if you can avoid actually burning the marijuana, you have probably eliminated the deleterious inhalants.
        This approach theoretically has been around for about 20 or 25 years, but there are problems with the existing instruments, which we have overcome. Unfortunately, we have to go from prototype to a reasonably manufacturable kind of thing, and that takes a while to develop. We also have some proprietary rights that we are currently wrapping up.
        Basically, the marijuana "joint" is a vaporization device and a decarboxylation device that uses a burning ember to super-heat an air stream. The THC does not come off the burning ember, but is distilled from the material in back of the ember. The ember itself is 600 degrees Celsius, which is probably three to four times too hot, but that is what you've got. When you have the delivery device being a little sheet of paper that you roll up around the material, that is pretty economical, and most people will go that way.
        However, we found that under milder conditions, you can also get a good transfer of cannabinoids. Now, if you combine this non-pyrolytic vaporization device with a Cannabis of extremely clean cannabinoid profile, then you have a combination of factors that let you deliver a high grade dose of a rather pure drug directly to the pulmonary system without the adverse effects possible with smoking marijuana.
        As an answer for my addendum to medical objection number three, what do you do about sterility? Well, basically there are three sorts of acceptable forms of sterilization, one of which we can implement, if demanded. There is the usual auto-claving, but that is not going to work out too well because everything gets soggy. There is ethylene oxide treatment, but that may cause chemical reactions because it is a pretty reactive molecule, and residues can hide in the plant crevices.
        Basically, we have "gone nuclear", as the expression has it. We can irradiate marijuana with a cobalt source and render it sterile of active organisms in the same manner in which imported spices are sterilized in the US. Personally, I am opposed to irradiating foods, but for very limited use, for medical use, this would seem reasonable. The medical field routinely sterilizes plastic syringes and other sorts of medical components. So, this is a well-worked-out technology.
        The advantages of inhaled cannabinoids are significant, we have seen that they are quickly available, allow accurate real-time titration, and there is no GI interference, that is, when you are vomiting or having diarrhea.
        Anandamide, I won't go into deeply because I am over my allowed time. However, it is a very interesting molecule. There is some potential for using this endogenous brain cannabinoid as an alternative to plant cannabinoids, at least in the ocular realm, where use of Cannabis has significant disadvantages. We think anandamides have advantages because they are not controlled substances, apparently penetrate corneal tissues nicely and we also happen to have a patent on them. However, some disadvantages are that they are chemically unstable and water insoluble.
        These problems have been overcome by the use of cyclodextrins. Anandamide has a chemical half life in water, what little will go into water, of about 12 hours. With its use in conjunction with cyclodextrins, which also increases anandamide's aqueous solubility a thousand-fold, this half-life is increased to six years. So, there is a dramatic increase in stability by putting them into cyclodextrins.
        I think that is it. Everyone can now go to lunch. Thank you. If you have questions and really aren't that hungry, I will be glad to field them.
        AUDIENCE PARTICIPANT: Does your vaporizer use heat?
        MR. PATE: Yes, you have to use some heat to get the cannabinoids volatized off, and additionally, in situ, the cannabinoid acids decarboxylate. The reason vaporization from the herbal material works so well, in contrast with the harshness of THC bronchial inhalers, is probably that the particle size of the aerosol is very fine due to its origin from a cannabinoid gland reservoir that has a diameter of only 75 to 100 microns to begin with. Also, the totaled surface area of all the plant's cannabinoid-laden glandular structures exposed to this hot air stream is huge, so evaporative transfer is quite efficient.
        DR. WATSON: Just one quick question for my personal interest. What is your background, your training?
        MR. PATE: I would say I have a rather checkered academic history. I started out with an interest in ethno-botany, sort-of from an anthropological standpoint, but then I traveled more toward plant biology, in which I have a Master's degree, because I became more interested in the role of the plant's secondary compounds in its ecology. From there, I became interested in the drugs that many of these secondary compounds are, so I went into pharmacognosy, but eventually dropped out of that program due to a "broken heart". Now at long last, because of an interest in changing those drugs to work better, I am almost finished with a doctorate in pharmaceutical chemistry. So, my path has been a little strange and certainly indirect!
        DR. WATSON: It sounds like a good match.
        DR. MUSTY: Dave, have you ever estimated what the production costs of THC would be, if you were doing it from a plant source, as compared with the cost of Marinol?
        MR. PATE: Our general reflexive response, based on some crude estimates, is that we can make THC cheaper than anyone can buy the precursors for the synthesis.

Resin glands

Resin glands on the surface of a Cannabis flower.