Abstract
Pancreatic cancer is the fourth leading cause of cancer death for both men and women in the United States. It is characterized by late diagnosis, poor prognosis and a lack of response to most conventional therapies and surgical resection is the only effective treatment for this dreadful disease. The five-year survival is less than 2% and the median survival after diagnosis is less than 6 months.|Accumulating evidence has linked pancreatic cancer growth with dietary fatty acid intake, especially intake of m-6 polyunsaturated fatty acids (PUFAs), such as iinoleic acid (LA) and arachidonic acid (AA), which enhance tumorigenesis through the production of eicosanoids. On the other hand, w-3 PUFAs, such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) that are found in high concentrations in fish oils, have anticancer effects. Two major metabolic pathways exist for arachidonic acid, namely the cyclooxygenase (COX) pathway and the lipoxygenase (LOX) pathway. Earlier investigations into the role of arachidonic acid metabolism in cancers focused mainly on the COX pathway, because of the epidemiological evidence that the incidence of colon cancer was greatly reduced in regular users of aspirin and other non-steroid anti-inflammatory drugs (NSAIDs). More recently, the role of the LOX pathway in human cancer has been more widely studied. Previous studies in our laboratory have shown that: (1) Both 5-LOX and 12-LOX mRNA and protein are over-expressed in human pancreatic cancer cells as well as pancreatic adenocarcinoma tissues. (2) The LOX products, 5-HETE and 12-HETE stimulate proliferation of pancreatic cancer cells through the activation of multiple intracellular signal pathways. (3) Blockade of the 5-LOX or the 12-LOX pathways inhibits proliferation and induces apoptosis in human pancreatic cancer cells.|Leukotrienes constitute a class of potent biological mediators of inflammation and anaphylaxis. Among them, leukotriene B4(LTB4) is one of the final products of the 5-LOX pathway of arachidonic acid metabolism and it has been widely implicated in the pathogenesis of several inflammatory diseases, such as asthma, psoriasis, rheumatoid arthritis and inflammatory bowel disease. However, its role in tumorigenesis has not been well studied. Role of the LTB„ metabolic pathway in the proliferation of human pancreatic cancer is the focus of this dissertation.|Using RT-PCR and immunoblotting, we confirmed that two G-protein coupled receptors for LTB4, BLT1 and BLT2 are expressed in several human pancreatic cancer cell lines. The identities of the receptors were confirmed by sequencing of the RT-PCR products. The sequences were identical to the reported sequences for BLT1 and BLT2 in GeneBank. Immunohistochemical assays demonstrated both BLT1 and BLT2 are overexpressed in human pancreatic adenocarcinoma tissues compared to normal pancreatic cancer tissues from multiple organ donors.|The effect of LTB, on the proliferation of human pancreatic cancer cells was investigated LTB, stimulated DNA synthesis and proliferation of pancreatic cancer cells. The production of LIB, in pancreatic cancer cells was increased upon EGF stimulation. LTB, also induced rapid and transient phosphorylation and activation of MEK and ERK1/2 kinases. The MEK inhibitors, PD98059 and U0126 as well as the selective LTB, receptor antagonist LY293111 all blocked LI B,-induced ERR 1/2 activation and cell proliferation. LTB, also induced phosphorylation and activation of p38 MAPK. The p38 MAPK inhibitor SB203580 was able to block the LTB,-induced activation of p38 MAPK but not cell proliferation. The serine/threonine kinase Akt/PKB was also phosphorylated following LTB, treatment. The PI-3 kinase inhibitor wortmannin partially blocked LTB,-induced Akt activation and cell proliferation. These results indicated that LTB, stimulated proliferation of pancreatic cancer cells is mediated at least in part, through the activation of MEK/ERK and PI-3 kinase/Akt cascades.|We next investigated the effect of abrogation of LTB, signal transduction on the proliferation of human pancreatic cancer cells. The selective LTB, receptor antagonist LY293111 inhibited DNA synthesis and cell proliferation in pancreatic cancer cells in vitro. LY293111 also induced marked apoptosis of these ceiis as measured by morphology, annexin V staining, F-actin staining and TUNEL assay. The molecular mechanisms of LY293111-induced apoptosis were further dissected. LY293111 induced the release of cytochrome c from the mitochondria into the cytosol, with the activation of caspase-9, caspase-3, caspase-7 and the cleavage of caspase substrate PARP. LY293111 also decreased the levels of the anti-apoptotic proteins Bcl-2, Mcl-1 and increased the level of pro- apoptotic protein bax. The distinctive S phase arrest induced by LY293111 involved the down- regulation of cyclin A, cyclin E and CDK2 levels, but the levels of p21 and p27 were not affected.|Based on the above in vitro results, we further investigated the effect of various LOX inhibitors and LY293111 on the growth of pancreatic cancer xenografts in athymic mouse models The non-selective LOX inhibitor NDGA, the selective 12-LOX inhibitor baicalein, as well as the selective LTB4 receptor antagonist LY293111, all markedly attenuated the growth of AsPC-1 and HPAC pancreatic cancer xenografts in athymic mice. These compounds also induced marked apoptosis of these tumor xenografts.|In conclusion, the current studies are the first reported studies on the importance of the LTB4 metabolic pathway in the proliferation of human pancreatic cancer. The findings on the role of LTB4 in promoting the proliferation of pancreatic cancer cells and the effect of inhibiting LTB4 signaling on the proliferation in these cells may provide a novel and promising therapeutic alternative for pancreatic cancer.