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12-Lipoxygenase (12-LOX) catalyzes the transformation of arachidonic acid into 12-hydroxyeicosatetraenoic acid (12-HETE). 12-LOX and its product 12-HETE have been implicated in cancer development and are a potential target for cancer chemoprevention. In our effort to study the effects of green tea polyphenols (GTPP) on arachidonic acid metabolism in the Min mouse, we investigated cyclooxygenase (COX) and lipoxygenase (LOX) activity in the microsomal and cytosolic fractions of the Min colon (stripped of all visible polyps) and Min colon polyps (MCT). COX was the major pathway for arachidonic acid metabolism in the colon with PGE2 being the major metabolite. Inhibition studies with NS398, a selective COX-2 inhibitor, suggested that COX-2 was responsible for 30% of the activity. On the other hand, PGE2 formation was not detectable in MCT. A metabolite that was only detectable in the MCT fractions coeluted with 12-HETE/8-HETE standards suggesting 12-LOX and/or 8-LOX activity. The majority of this activity was located in the microsomal fraction. The general LOX inhibitor nordihydroguaiaretic acid (25 μM) completely inhibited activity in the microsomal and cytosolic MCT fractions. Baicalein (5 μM), a selective 12-LOX inhibitor completely inhibited the cytosolic 12-LOX (c12-LOX) activity and reduced the microsomal 12-LOX (m12-LOX) activity by 75%, thereby suggesting 12-LOX activity. However, the microsomal and cytosolic fractions reacted differently to the presence of glutathione. Glutathione (1mM) increased m12-LOX activity by 2-fold while c12-LOX activity was completely inhibited. The 2 fractions also differed in their response to the presence of GTPPs. Epicatechin-3-gallate (ECG) and epigallocatechin-3-gallate (EGCG) were the most effective GTPPs at inhibiting both enzymes. IC50 values for m12-LOX were 5 μM and 8 μM while IC50 values for c12-LOX were 10 μM and 40 μM for ECG and EGCG, respectively. Moreover, EGCG had a complex effect on the c12-LOX. Low concentrations of EGCG increased c12-LOX activity reaching a maximum when EGCG was present at a concentration of 5 μM after which a dose dependent inhibition was observed. The above differences between the c12-LOX and m12-LOX suggest that 2 functionally distinct forms of 12-LOX may be present in the colon tumor tissue. The results of this study demonstrate the presence of 12-LOX activity in colon polyps resulting in a shift in arachidonic acid metabolism from the COX pathway in the ‘tumor free’ colon to the 12-LOX pathway in the colon tumors and inhibition of this activity by GTPP suggests that green tea may exert it chemopreventive effects through inhibition of 12-LOX. This study was supported by NIH grant CA096694.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]