Monoterpenes

Monoterpenes

"Monoterpenes appear to act through multiple mechanisms in the prevention and chemotherapy of cancer. Several researchers are investigating these mechanisms and finding that, although the exact mechanism was not what they had assumed, the monoterpenes, limonene, and perillyl alcohol [and perillic acid and geraniol] have a profound antitumor activity on pancreatic cancer (Elson et al. 1994; Gelb et al. 1995; Crowell et al. 1996; Gould 1997; Bardon et al. 1998; Crowell 1999)." See: Limonene, Perillyl Alcohol and Geraniol


Monoterpenes in Cancer Prevention and Therapy

Abstract

When people first hear the word monoterpene (MT), typically the "terpene" portion conjures up images of some sort of cleaning fluid such as turpentine. Associated with this image is the idea that they are poisons. While the MT's are indeed used as cleaning agents due to their solvent properties, they are far from being poisons. Indeed, nothing could be further from the truth for these naturally occurring and health-promoting substances. While the solvent properties of monoterpenes have been exploited clinically to dissolve gallstones,[ 1] the monoterpenes are also the focus of much investigation in the areas of both cancer prevention and therapy. The anticancer properties of the MT's are discussed in the context of clinical data and future directions are explored.

Introduction

Cancer prevention, inhibition, and regression are the most noteworthy attributes of the MT's. D-limonene (DL) and perillyl alcohol (POH) have been shown to be chemopreventive against mammary,[ 2] liver, lung, UV-induced skin cancer[ 3] and chemotherapeutic against both experimental mammary and pancreatic tumors. Perillyl alcohol stands out as effective against human pancreatic cancer,[ 4] colon, liver[ 5] to reduce vein graft intimal hyperplasia,[ 6] as chemopreventive against colon carcinogenesis, prostate and lung cancer.[ 7] Several of these are discussed below in the context of the clinical data.

The Monoterpenes

As seen in figure 1, DL is a monocyclic MT with POH a metabolite of DL, being its hydroxylated form. There are a number of synonyms for limonene, include the following: 1,8( 9)-p-Menthadiene; 1-methyl-4-( 1-methylethenyl) cycloclohexene; 1-methyl-4-isopropenyl-1-cyclo-hexene; alpha-limonene; dipentene; limonene; p-mentha-1,8-diene. They are found in essential oils of many plants including lemons, oranges, grapefruit, caraway, dill, bergamot, peppermint, spearmint, grasses and tomatoes. They are also associated with vegetables and some evergreen trees.[ 8] POH is often distilled from lavender, found in cherries, mint, celery seeds[ 9] and can be produced synthetically. It is typically used as flavoring agents, food additive, and fragrance and has been found to be a major volatile component of mother's milk.[ 8]

Figure 1: structures of Perillyl Alcohol and Limonene

D-limonene has different metabolites for different animals. In humans, the three major metabolites after an oral dosage are perillic acid, dihydroperillic acid, and limonene-1,2-diol. It is thought that the metabolic precursors of the first two are perillyl alcohol and perillyl aldehyde.[ 9] Moreover, many people regularly consume DL everyday without even knowing it. This is because DL is found in things such as orange juice at concentrations ranging from 10-100 ppm and chewing gum, which contains up to 2,300 ppm.[ 8]

How do they work?

In order to understand the mode of action of the anticancer properties of the monoterpenes, it is necessary to review some of the molecular events that surround cancer. Normal, non-cancer cells live a limited and constrained life. Once normal cells are formed, they have a finite number of cell divisions they undergo during their life-span and, with few exceptions, they remain relatively localized to the same point in the body during this entire process. As a cell lives and ages, it maintains an "awareness" of its surroundings via cell-cell communication molecules both on the cell surface and secreted. This allows the cells to determine if all is well and good and instructs them to maintain their relative positions in the body. Towards the end of a cell's designated life-span, certain cellular events occur which instruct the cell to terminate. This is called "apoptosis" or programmed cell death and is a very normal and useful event for clearing away the old and making room for the new. While it may not sound like the most compassionate thing to do to old cells, the evolutionarily-derived utility of such action affords the body a mechanism to remove cells which might be on their way to becoming cancer cells via mutations.

All cells suffer mutations. These mutations can come as a result of environmental agents, toxic products of metabolism, or arise spontaneously. The exact mechanism is not important. What is important, however, is that evolution has built into the cells, mechanisms to deal with these. Apoptosis is one way to get rid of cells which have had ever-increasing chances of suffering mutations that are irreparable. The cellular machinery is such that the basal level of mutations are usually successfully corrected by processes such as Nucleotide Excision Repair (NER).[ 10] When, for various reasons, they cannot be, then generally, apoptosis ultimately will make the mutations moot.

There are several stages in the formation of cancer cells. The first is the initiation stage. During initiation, some mutation has to occur that results in either a loss of function (exemplified by p53, tumor suppressor gene) or a gain of function (e.g., virally derived oncogenes in which there is production of aberrant proteins having normal cellular counter-parts). The key --mutation needs to occur and not be corrected by the usual cellular mechanism. At this point, cells will typically become described as "immortalized." When this occurs, the cells no longer have finite life-spans, but can live indefinitely. This allows the cancer cells to escape the process of apoptosis. At this stage, the cancer cell is still relatively harmless.

The next stage is the promotion/progression stage of cancer development. Now the cancer cell, usually through a result of even more mutations, becomes "transformed" and no longer acts like a normal cell. The transformed cell is now unrestricted in both its life-span and localization. As stated above, a normal cell will sense its environment and be restricted in its growth. The transformed cell no longer abides by such rules and will multiply, literally piling up on top of its neighbors. Typically, there is an associated perturbation of the cellular glycosylation pattern.[ 11, 12] Also, the morphological structure of the cell itself changes, from being flat with multiple projections, to being much smaller and rounded up.

The final stage is metastasis. Here, the cancer cells have progressed to the point where they are no longer localized to one ever-growing tumor. They gain the ability to enter the blood vessels (intravasation), move to a different location in the body, stop and exit the blood vessel (extravasation) and establish a new site of tumor development. Eventually, the body becomes fiddled with tumors consuming the bulk of available energy. The body is literally eaten alive as evidenced by the cancer-associated cachexia (body wasting).

In that dreadfully bleak picture of cancer development, there are some shining points of lights. The MT's, found in essential oils, are such points of light, offering some hope in the struggle to prevent and treat cancer. The best news is that DL and POH can not only prevent but also treat cancer. That is to say, the MT's can act before a cancer is established and in the cases where cancer is already present, they can cause a regression of the tumor, oftentimes completely. In fact, levels of tumor regression as high as 81% have been achieved for small mammary carcinomas and up to 75% for advanced mammary carcinomas.[ 22] They do this in at least six ways. First, during the initiation phase of carcinogenesis they induce (cause the body to make more of) Phase I and II carcinogen-metabolizing enzymes, resulting in carcinogen detoxification.[ 13] An example of such a Phase II enzyme would be glutathione S-transferase.[ 14] Second, post-initiation phase, they have been shown to increase cell redifferentiation. This causes the potential cancer cells to take on a more normal morphology. Third, they can induce apoptosis in otherwise immortalized (see above) cells. Fourth, they have been shown to inhibit the isoprenylation[ 15] of the cellular products (Ras[ 16]) of oncogenes. Simply put, the proteins from oncogenes, which on the whole are cell-growth regulating proteins, need to be modified (referred to as "post-translational modification") by a process called prenylation in order to be placed in a membrane where they are active. If the proteins from oncogenes do not undergo isoprenylation, they do not cause the cell to behave as a cancer cell and hence cancer inhibition results. It is believed that this prenylation inhibition occurs at both the farnesyl and geranylgeranyl transferases. Fifth, MT's have been shown to inhibit the conversion of lathesterol to cholesterol, resulting in positive therapeutic results due to the fact that many tumor cells are deficient in oxidative phosphorylation and use glycolysis as the sole energy source.[ 22] Sixth, the MT's have been shown to enhance activation of inhibitory growth factors such as TGF[Beta], which has been suggested to potentially inhibit br east cancer cells.[ 23]

Clinical data

Because the metabolites of monoterpenes, which vary from species to species, also show anticancer effects, it is necessary to discuss dosage in the context of the specifics of each study.

In rats treated with DMBA (an agent that induces tumor formation) POH and DL had complete regression rates of 81 and 68%. Interestingly, the amount of DL required to achieve these results was four times higher than POH.[ 17, 18] The dosage for mammals in vitro is suggested to be approximately 2.5g/kg for POH (roughly five times that for DL). The mammalian in vivo data suggests a dose of 7.5 g/m2. In humans, for POH, this would translate to approximately 10-15 g/day. While most of the work cited above has been done in mammals, human clinical trials have been reported.
Human phase I clinical results for POH used in the treatment of advanced malignancies in humans, have been reported.[ 19] Dosages ranging from 800 mg/m2/dose to 2400 mg/m2/dose were assayed for tolerability. The main toxicity was gastrointestinal and included nausea and vomiting, anorexia, unpleasant taste, satiety, and eructation. The main metabolites were perillic acid and dihydroperillic acid. The authors reported evidence of POH efficacy in metastic colorectal cancer where one patient out of two colorectal cancer patients, of the 15 total evaluable patients in this safety trial, was free of tumor. Three other patients in this study had their advanced stage disease (two prostate cancer patients and one patient with adenocystic carcinoma of the salivary gland) stabilized. While the conclusions were not as positive as hoped for, there was some encouragement. The disease was shown to stabilize for 6 months, although no objective tumor responses were observed. The authors concluded that the dosage interval should be increased and the results of that have not been announced. It should be noted that these were advanced malignancies, which had been refractory to prior treatments. In other words, this could be viewed as a worst case scenario. Also, the use of 2,400 mg/m2/dose is less than the recommended dosage.[ 18]

A telephone conversation with the Principle Investigator of the above study, revealed that resulting Phase II studies are currently ongoing for Breast and Prostate cancer using POH. Additionally, because of problems with the original NCI formulation (half soy bean oil and half POH in 250 mg capsules) there are currently repeated Phase I studies ongoing with a new formulation of 90% POH in 700 mg capsules. Additional problems with the original formulation were the complications that any phytoestrogens present in the soybean oil, might contribute to the interpretation of the data. However, the results look promising, in that an effect was seen for both breast and prostate cancer.

The main study, using DL, has been reported by a British group who saw disease stabilization for both breast and colon cancer patients using 8g/m2/day.[ 20] Presently, there are no other ongoing studies for DL, though the National Cancer Institute in the US is said to be considering it. One of the problems with getting a study going for prevention is that large, randomized studies for five or six years need to be performed. It is difficult to get companies interested in doing those, despite the exciting data, because of the huge investment of time and money coupled with the difficulty of patenting the naturally occurring compounds.

Toxicology

The bulk of the toxicity data for the monoterpenes comes from the study of limonene in animals and has been well presented by Von Burg.[ 8] To briefly summarize, limonene can cause skin sensitization in susceptible people. A blood clearance rate of 1.11/kg/hr has been established with the majority being eliminated through the urine. Immunosuppression has been observed in mice. This is particularly interesting because aberrations of the immune system have been postulated for causes of cancer cell proliferation, which seems counter-intuitive. There appears to be no genotoxicity (Ames test) though straight orange peel oil was found to have slight promotional activity for mouse skin tumors initiated with (DMBA). However, the promotional activity was not observed when either limonene or straight orange peel oil were added to the diet, suggesting a relationship between promotion and irritant properties of the straight oil. Similarly, there appears to be no neurotoxicity. It should be noted t hat at relatively high dosages (2363 mg kg-1) orally administered to mice, decreased body weight and increased bone abnormalities were observed in fetuses. However, at lower dosages (591 mg kg-1), no effects were seen, either maternally or in the fetus. Theoretically, there is a possible lethal oral dose for humans in the range of 0.5 - 5 g/Kg.

Current sources

As stated above, the MTs are found readily occurring in nature. However, it is probable that supplementation is necessary to achieve the positive therapeutic and preventive indices. Currently, POH use is restricted in the US due to the issuance of an Investigational New Drug (IND) application. Nevertheless, there are major laboratory chemical supply companies, which do carry POH as a chemical reagent, though not for human consumption.

At this time, there are very few products on the European and US markets of which the author is aware. Very few companies actually have encapsulated, or tableted MT products.[ 21] However, it is possible to purchase a highly purified (>=95%) DL product from the larger flavor companies and is relatively inexpensive. The caveat being that they have large minimum quantities (typically over 25 lbs) required for purchase. However, to achieve the dosages suggested in the literature, these quantities would be required anyway and the actual cost is minimal.

With the high level of impressive scientific data available on the efficaciousness and clinical tolerability of these naturally occurring nutrients, the monoterpenes are positioned to take a strong role in the war on cancer. They offer a strong ally for the more established routines of cancer therapy and deserve to be given attention. The next few years will be an exciting time as more and more of the Phase II clinical trials are completed and results reported. While less than hoped for results have at times been reported,[ 23] it should be kept in mind that the studies were done on advanced stage carcinomas. It is not unreasonable to expect more favorable results with earlier treatment in the course of the disease. Also, despite the reluctance of major companies to fund the clinical studies on preventive properties of MT's. there will inevitably be a continued stream of anecdotal support for their efficacious chemopreventive usage. The public can expect to see new products. such as DL combined with Red Rice Yeast, which contains lovastatin, whose chemopreventive properties[ 5] may be additive or synergistic to those of the MT's alone. Until then, enjoy your fruits and vegetables!

Correspondence:

Mark A. Brudnak, PhD, ND MAK Wood, Inc. 1235 Dakota Drive, Units E-F Grafton, Wisconsin 53024-9429 USA 262-387-1200 (3#) / Fax 262-387-1400 Email: makwood@earthlink.net

References

1. Igimi, H., Hisatsuga, T., and Nishimura, M. The use of d-limonene as a dissolving agent in gallstones. Dig. Dis., 2 1:926-939, 1976.

2. Crowell, PL. Monoterpenes in breast cancer chemoprevention. Breast Cancer Research and Treatment 46:191-197, 1997.

3. Uedo N, Tatsuta M, Iishi H, Baba M, Sakai N, YanoH, Otani T,. Inhibition by D-limonene of gastric carcinogenesis induced by N-methyl-N'-nitro-N-nitroguanidine in Wistar rats. Cancer Letters 137(2):131-6 1999.

4. Stark MJ, Burke YD, McKinzie JH, Ayoubi AS, Crowell PL. Chemotherapy of pancreatic cancer with the monoterpene perillylalcohol. Cancer Lett 9:15-21.1995.

5. Broitman SA, Wilkinson J 4th, Cerda S. Effects of Monoterpenes and Mevinolin on Murine Colon Tumor CT-26 In Viro and its Hepatic "Metastases" In Vivo. Adv Exp Med Biol 401:111-30. 1996.

6. Fulton GJ, Barber L, Svendsen E, Hagen PO, Davies MG.Oral monoterpene therapy (perillyl alcohol) reduces vein graft intimal hyperplasia. J Surg Res Apr;69(1):128-34. 1997.

7. Reddy B.S., Wang C.X., Samaha H., Lubet R. Steele V.E., Kelloff G.J., Rao C.V. Chemoprevention of colon carcinogenesis by dietary perillyl alcohol. Cancer Res Feb. 1997.

8. Von Burg, R. Limonene Journal of Applied Toxicology 15 (6):495-9 Nov-Dec. 1995
9. Hohl, RJ Monoterpenes as Regulators of Malignant Cell Proliferation. Advances in Experimental Medicine and Biology 401:137-46. 1996.

10. Lindahi T and Wood Rd Quality Control by DNA Repair. Science 286 (5446):1897-905 Dec 3. 1999.

11. Dennis JW, Granovsky M, Warren CE. Glycoprotein glycosylation and cancer progression. Biochimica et BiophysicaActa 1473:21-34.1999.

12. Taniguchi, et al. Implications of N-acetylglucosaminyltransferases III and V in cancer: gene regulation and signaling mechanism. BiochimicaEtBiophysica Acta 1455;287-300. 1999.

13. Kensler TW Chemoprevention by Inducers of Carcinogen Detoxication Enzymes. Environmental Health Perspectives 105. Suppl. 4 June 965-970. 1997.

14. van Lieshout, EMM, Bedaf MMG, Pieter M, Ekkel C, Nijhoff WA and Peters WHM. Effects of dietary anticarcinogens on rat gastrointestinal glutathione S-transferase theta 1-1 levels. Carcinogenesis19 (11) 2055-2057. 1998.

15. Galli I, Uchiyama M, Wang TSF. DNA Replication and Order of Cell Cycle Events: A Role for Protein Isoprenylation? Biol. Chem, 378 963-973 1997.

16. Yamamoto T, Taya S, Kaibuchi K. Ras-Induced Transformation and Signaling Pathway. J. Biochem. 126, 799-803. 1999.

17. Gould MN, Crowell PL, Elson CE, Ren Z. Uses of Perillic Acid Methyl Ester. United States Patent No. 5,470,877. 1995.

18. Gould, MN, Crowell PL, Elson CE. Regression of Mammalian Carcinomas. United States Patent No. 5,414,019. 1995.

19. Ripple GH, Gould MN, Stewart JA, Tutsch KD, Arzoomanian RZ, Alberti D, Feierabend C, Pomplun M, Wilding G, Bailey HH. Phase I clinical trial of perillyl alcohol administered daily. Clin Cancer Res May;4(5):1159-64. 1998.
20. Vigushin DM, Pooh Gk, Boddy A, English J, halbert GW, Pagonis C, Jarman M, Coombes RC. Phase I and pharmacokinetic study of D-limonene in patients with advanced cancer. Cancer Research Campaign Phase I/II Clinical Trials Committee. Cancer Chemother. Pharmacol. 42(2):111-7. 1998.

21 Solgar Vitamin and Herb Company. http://www.solgar.com.
22. Haag JD and Gould MN Mammary carcinoma regression induced by perillyl alcohol, a hydroxylated analog of limonene. Cancer Chemother. Pharmacol. 34:477-483. 1994.

23 Jirtle RL, Haag JD, Ariazi EA, Gould MN. Increased mannose 6-phosphate/insulin-like growthfactor II receptor and TGF-[Beta] 1 levels during monoterpene-induced regression of mammary tumors. Cancer Res 53:3849. 1993.
Previously published in Positive Health issue #53.
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By Mark A. Brudnak

See: http://cancer.lef.org/pancreatic-3.html

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