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Lipoxygenase and Cancer: Incriminating Evidence For Dietary Arachidonic and Linoleic Acids
James Meschino DC, MS, ROHP
Mounting evidence suggests that lipoxygenase (LO)-catalyzed products have a significant influence on the development and progression of human cancers. Compared with normal tissues, significantly elevated levels of LO metabolites have been found in lung, prostate, breast, colon, and skin cancer cells, as well as in cells from patients with both acute and chronic leukemias. LO-mediated products trigger diverse biological activities required for cancer growth, and influence growth factor and transcription factor activation, oncogene induction, stimulate tumor cell adhesion, and regulate apoptosis.
During the past 10 years, pharmacological agents that specifically inhibit the LO-mediated signaling pathways are now commercially available to treat inflammatory diseases such as asthma, arthritis, and psoriasis. LO inhibitors are now being explored as agents to prevent and treat certain cancers.
Mechanisms Through Which Arachidonic Acid and Leukotrienes Encourage Cancer Development And Growth
Eicosanoids derived from the arachidonic acid cascade have been implicated in the pathogenesis of a variety of human diseases, including cancer, and appera to play important roles in tumor promotion, progression, and metastatic disease. Although most attention has focused on PG2 and other COX-derived metabolites, mounting evidence suggests that LO-catalyzed products, LTs, and HETEs also exert profound biological effects on the development and progression of human cancers. For example, 12-LO mRNA expression has been well-documented in many types of solid tumor cells, including those of prostate, colon, and epidermoid carcinoma. Also, 12(S)-HETE production by some tumor cells, including prostate cells, has been positively correlated to their metastatic potential.
Studies indicate that 12(S)-HETE is a critical intracellular signaling molecule, stimulating PKC and eliciting the biological actions of many growth factors and cytokines that regulate transcription factor activation and induction of oncogenes or other gene products needed for cancer growth. Some of these include EGF, FGF, PDGF, tumor necrosis factor, granulocyte-macrophage colony-stimulating factor, and both IL-1 and IL-3. Additionally, PKC activation by 12(S)-HETE mediates the release and secretion of cathepsin B, which is involved in tumor metastasis and invasion, particularly in colon cancer cells.
As well, tumor cell synthesis of 12(S)-HETE stimulates adhesion by increasing the surface expression of integrin receptors. Besides 12(S)-HETE, other LO metabolites, particularly the 5-LO products (5-HETEs), have been implicated in cancer development. In fact, studies show that 5-HETE directly stimulates prostate cancer cell growth.
In general, evidence from studies in human cancer cells shows that the LO pathways are involved in carcinogenesis in several major tissues. More importantly, a few studies in animal carcinogenesis models show that inhibition of the LO pathways also may inhibit carcinogenesis
Two naturally occurring agents, Baicalein and esculetin are among several experimental compounds demonstrating 12-LO inhibitory activity that may show promise as antiproliferative agents. (1) These agents will be discussed in the next section
Lipoxygenase and Prostate Cancer
Diets high in fat are associated with an increased risk of prostate cancer, although the molecular mechanism is still unknown. (2) The increased risk of prostate cancer associated with high dietary intake of animal fats shown in multiple population-based studies may in part be explained by the rich sources of fatty acids like arachidonate in these diets that favor conversion to proinflammatory eicosanoids. (4)
Studies have previously reported that arachidonic acid, an omega-6 fatty acid common in the Western diet, stimulates proliferation of prostate cancer cells through production of the 5-lipoxygenase metabolite, 5-HETE (5-hydroxyeicosatetraenoic acid). Research reveals that 5-HETE is also a potent survival factor for human prostate cancer cells. These cells constitutively produce 5-HETE in serum-free medium with no added stimulus. Exogenous arachidonic acid markedly increases the production of 5-HETE. Inhibition of 5-lipoxygenase by MK886 completely blocks 5-HETE production and induces massive apoptosis in both hormone-responsive (LNCaP) and -nonresponsive (PC3) human prostate cancer cells. Exogenous 5-HETE protects these cells from apoptosis induced by 5-lipoxygenase inhibitors, confirming a critical role of 5-lipoxygenase activity in the survival of these cells. These findings provide a possible molecular mechanism by which dietary fat may influence the progression of prostate cancer. (2)
Altered eicosanoid biosynthesis in relation to prostate cancer development has been well documented. Initial studies found dramatically reduced levels of arachidonic acid; 10-fold greater turnover in malignant compared to benign prostatic tissue suggested a possible increase in metabolism via the LO and COX pathways in this tissues. Linoleic acid stimulated cell growth in experiments conducted in human prostate cancer cells, whereas indomethacin, esculetin, and piroxicam inhibited it, which provides significant proof of the involvement of eicosanoids in prostate cancer cell proliferation. Although attention has focused on COX-derived products, particularly PGE2, COX inhibitors indomethacin and aspirin failed to reduce human prostate PC3 cell DNA synthesis, although arachidonic acid antagonist eicosatetraynoic acid did reduce synthesis, suggesting that LO products are essential in modulating prostate DNA synthesis. Thus, researchers now recognize that inhibiting or down-regulating the LO synthesis pathway is an important target for prevention and management of certain cancer, including prostate cancer.
Studies show that 5-HETE, particularly the 5-oxo-eicosatetraenoic form, stimulates PC-3 cell growth similarly to arachidonic acid. 5-HETEs also effectively reversed growth inhibition produced by a drug known as MK-886 (Merck), a FLAP inhibitor
In addition to 5-LO products, the 12-LO metabolite 12(S)-HETE has been shown to play a critical role in prostate tumor metastasis and invasion . In 122 matched normal and cancerous prostate tissues, 12-LO mRNA expression was confined to prostate epithelial cells and elevated in malignant cells. As well, elevated levels of 12(S)-HETE mRNA significantly correlated with advanced stage, poor differentiation, and invasive potential of prostate cancer cells. Indeed, addition of 12(S)-HETE to rat Dunning R3327 and human prostate adenocarcinoma cells significantly increased their cellular motility and ability to invade basement membrane and increased their metastatic activity.
Additional biochemical evidence demonstrated that 12(S)-HETE stimulates secretion of cathepsin B, which is involved in tumor metastases and integrin expression in other tumor cells, providing support for the importance of LO products in the development and spread of prostate and other human cancers. It has also been shown that 12(S)-HETE involves selective activation of membrane-associated PKC, which is known to be involved in cancer growth. (1) This information takes on greater significance with the appreciation that cells that have the appearance of prostate cancer can be found in the prostate gland of nearly half of all men over the age of 50. (4)
Lipoxygenase and Breast Cancer
Studies have shown that diets rich in linoleic acid, stimulate mammary tumor growth induced by either MNU or DMBA, two frequently used carcinogens in animal used in animal studies. Conversely, dietary supplementation with n-3 polyunsaturated fatty acid, particularly fish oils, has been shown to retard mammary tumor growth and metastasis in a variety of animal tumor models. Several studies have shown that mammary tissue of rats fed 20% menhaden oil produced lower levels of LTB4 and PGE2 compared with groups receiving 20% corn oil or primrose oil, supporting the theory that PGs and LTs may be involved in mammary carcinogenesis
Markedly elevated levels of both COX and LO metabolites have been documented in human breast cancer tissue compared with tissue from patients with benign disease.
Other studies have indicated that linoleic acid enhanced the invasive capacity of breast cancer cells and that this effect could be completely blocked by adding esculetin, an inhibitor of 5- and 12-LO, but not by adding the COX-specific inhibitor piroxicam. This evidence strongly implicates the lipoxygenase pathway in cancer development.
The interaction of growth factors, such as epidermal growth factor (EGF) with their receptors, on breast cancer cells can lead to the hydrolysis of phospholipids and release of fatty acids, such as arachidonic acid, which can be further metabolized by the lipoxygenase (LO) pathway. Several LO products have been shown to stimulate oncogenes and have mitogenic and chemotactic effects.
More specifically, LO products, such as the hydroxyeicosatetraenoic acids (HETEs), have been shown to have actions highly relevant to cellular growth and migration. They have significant mitogenic and chemotactic effects and also can stimulate the expression of several oncogenes and other studies suggest that the biosynthesis of 12(S)-HETE by tumor cells is a determinant of their metastatic potential. The LO products of linoleic acid have been shown to potentiate the mitogenic effects of epidermal growth factor and linoleic acid can stimulate the growth of MCF-7 breast cancer cells.
There is also evidence to show that leukocyte 12-LO mRNA expression is upregulated in breast cancer cells and tissues, compared with their normal counterparts
Results of other studies also suggest that activation of the 12-LO pathway may play a key role in basal and EGF-induced breast cancer cell growth. As such, it appears that 12 LO end products up-regulate epidermal growth factor receptors, which are known to be involved in breast cancer development. In fact, treatment with EGF, a potent breast cell growth factor, caused a significant increase in 12-LO enzyme activity, as well as leukocyte-type 12-LO protein expression in MCF-7 cells. These results suggest that 12-LO expression is enhanced in breast cancer, and this increased expression may be secondary to locally derived growth factors involved in breast cancer cell growth and development. This is further supported by the specific inhibition of the growth of MCF-7 cells by LO inhibitors, but not CO, inhibitors. (3)
Lipoxygenase And Other Cancers
12- and 15-HETE are major arachidonic acid metabolites have also been incriminated in squamous epithelial carcinomas of the head and neck, Also, 12(S)-HETE is the predominant metabolic product of metastatic B16 melanoma cells and excess LT production, specifically LTC4, has been documented in cells from patients with both acute and chronic leukemias
Agents Shown To Block The LO and CO Pathways And Inhibit Cancer
Non Steroidal Anti-Inflammatory Drugs – Among the tumor types that have been shown to overexpress COX-2 is prostatic adenocarcinoma. Both selective and nonselective COX-2 inhibitors, such as celecoxib and sulindac, respectively, have been shown to be effective in preventing the progression of premalignant colorectal adenomas in preclinical and clinical studies as well as in preclinical studies in bladder, skin, and mammary gland. It has been suggested that these NSAIDs may be useful for the prevention or therapy of prostate cancer as well. Indeed, both sulindac and celecoxib have been shown to inhibit the growth or induce apoptosis in human prostate cancer cell lines in vitro, and the NSAID flufenamic acid has been shown to inhibit the expression of the androgen receptor in LNCaP cells. Recently, an inverse association between aspirin or ibuprofen use and prostate cancer development was noted in several case control studies. (4) Unfortunatley, the long-term use of NSAIDs is associated with gastrointestinal erosion, ulcers, intestinal bleeding, as well as liver and kidney toxicity. As such, their regular use as prophylactic cancer intervention may produce serious and life-threatening side effects and are clearly contra-indicated in patients with certain co-morbidity issues (e.g. previous intestinal ulcers, kidney disease/dysfunction/transplant etc) (5,6,7)
Flavonoids (e.g. Baicalein), Esculetin, Curcumin and Tea Polyphenols – A number of natural occurring agents that affect arachidonate metabolism, either via cyclooxygenase or the alternative 5-lipoxygenase have demonstrated chemoprevention effects against cancer. Some of these include flavonoids, curcumin, and tea polyphenols. (4) Others include esculetin and hydroxamic acids.
Esculetin and Baicalein, two naturally occurring agents, have been shown to inhibit lipoxygenase enzymes and modulate cancer development and growth. In vivo studies have shown that esculetin significantly inhibits the development of mammary tumors induced by DMBA in female Sprague Dawley rats fed either a high-fat (20% soybean oil) or low-fat (0.5% soybean oil) diet. As well, esculetin has been effective in reducing the invasive and metastatic activity of malignant tumor cells. Murine melanoma cells pretreated with 50 µM esculetin and injected into syngeneic mice produced significantly lower numbers of melanoma colonies on the lung surface and fewer tumor metastases. Baicalein, a flavonoid isolated from Scutellaria baicalensis Georgy roots (Chinese scullpcap), is a key component of Japanese herbal medicine Sho-saiko-to, commonly used to treat chronic liver diseases in Japan. Early studies conducted in rat platelets showed that low concentrations of baicalein exert potent 12-LO inhibitory activity, whereas much higher levels are required for COX inhibition. Other studies suggest that baicalein also blocks 5-LO and production of LTC4.
The antiproliferative effects of baicalein have been reported in several in vitro models. One study showed that baicalein reduced DNA synthesis and inhibited growth in two human hepatoma cell lines. Similar growth-suppressive effects of baicalein have been reported by other investigators in various hepatocellular cancer cell lines. Likewise, treating human breast cancer cells (MCF-7) with baicalen at microgram levels produced potent antiproliferative activity. In human T-lymphoid leukemia cells (CEM) stimulated with 10% FCS, showed reduced cell proliferation with addition of baicalein in a dose dependent fashion, with evidence of reduced activity of messengers known to be of significance in cancer growth. Rat studies showed that baicalein induced apoptosis in carcinosarcoma cells in a dose-dependent manner. Nevertheless, these initial data suggest that baicalein may be an effective chemopreventive agents.
Hydroxamic acids have also shown lipoxygenase inhibition and reduced cancer growth. N-benzyl-N-hydroxy-5-phenylpentanamide (BHPP), one of a series of hydroxamic acids, originally developed by Rorer, that was found to exert 5-LO inhibitory activity in rat leukocytes. Additional studies demonstrated 12-LO inhibitory activity in Lewis lung carcinoma cells and porcine leukocytes and in murine B16a melanoma cells. Of interest to natural health practitioners is the fact that certain hydroxamic acid are naturally occurring in cernitin pollen extract, often used in the treatment of prostate enlargement. A cernitin-derived hydroxamic acid has been shown to inhibit replication of certain human prostate cancer cells under experimental conditions. (Fouad K. et al, Prostate. 2006)
Many other experimental studies have provided evidence for the cancer inhibition effects of BHPP via its inhibitory influences on the 5 and 12 lipoxygenase enzymes. (1)
In summary, evidence suggests that tumor cells and several normal cells have LO activity, and both arachidonic acid and linoleic acids are converted to LO products, such as HETEs and hydroxyoctadecadienoic acids (HODEs). The 12-LO product, 12(S)-HETE, has been shown to play an important role in the metastatic process. 12(S)-HETE mediates the adhesion of tumor cells to the subendothelial matrix after endothelial retraction by a protein kinase C-dependent process. LO products, such as 12- and 15-HETE, also have mitogenic effects on endothelial cells.
Research demonstrates that 12-HETE is the predominant arachidonic acid metabolite produced by highly metastatic tumor cells. Furthermore, these highly metastatic cells synthesize much greater amounts of 12-HETE than the low metastatic tumor cells. Thus, an increased concentration of 12-HETE, produced by activated platelets, the tumor cells themselves, leukocytes, or by vascular cells, are thought to be affiliated with the proliferative and metastatic processes
LO metabolites of arachidonic acid have been reported to mediate tumor necrosis factor-induced protooncogene c-fos expression . In addition, EGF-induced mitogenic activity has been linked to the formation of LO products of linoleic acid, the HODEs
Thus, a growing body of evidence suggests that specific metabolites of arachidonic and/or linoleic acid serve as central elements in signal pathways necessary for cell mitogenesis, as induced by growth factors or oncogenic transformation. (3)
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